Patent application title: IgG Fc Variants for Veterinary Use
Inventors:
IPC8 Class: AC07K1628FI
USPC Class:
1 1
Class name:
Publication date: 2020-11-19
Patent application number: 20200362034
Abstract:
Provided are various embodiments relating to variant IgG Fc polypeptides
of companion animals having increased Protein A binding for ease of
purification, decreased C1q binding for reduced complement-mediated
immune responses, decreased CD16 binding (e.g., for reduced
antibody-dependent cellular cytotoxicity (ADCC) induction, increased
stability, and/or the ability to form heterodimeric proteins. In
addition, various embodiments relating to antibodies and fusion proteins
comprising such variant IgG Fc polypeptides are provided. Also provided
are various embodiments relating to contiguous polypeptides comprising
one or more variant GLP1 polypeptide(s) having improved serum half-life.
Further provided are various embodiments relating to contiguous
polypeptides or heterodimeric polypeptides comprising a GLP1 polypeptide
and a glucagon polypeptide as a dual GLP1 receptor and glucagon receptor
agonist. In various embodiments, such polypeptides may be used to treat,
for example, diabetes, obesity, or related indications, in companion
animals, such as canines, felines, and equines.Claims:
1. A polypeptide comprising a variant IgG Fc polypeptide comprising at
least one amino acid modification relative to a wild-type IgG Fc
polypeptide of a companion animal species, wherein the variant IgG Fc
polypeptide has increased binding affinity to Protein A relative to the
wild-type IgG Fc polypeptide.
2. A polypeptide comprising a variant IgG Fc polypeptide comprising at least one amino acid modification relative to a wild-type IgG Fc polypeptide of a companion animal species, wherein the variant IgG Fc polypeptide has reduced binding affinity to C1q and/or CD16 relative to the wild-type IgG Fc polypeptide.
3. The polypeptide of claim 1 or claim 2, wherein the variant IgG Fc polypeptide binds to C1q and/or CD16 with a dissociation constant (K.sub.d) of greater than 5.times.10.sup.-6 M, greater than 1.times.10.sup.-5 M, greater than 5.times.10.sup.-5 M, greater than 1.times.10.sup.-4 M, greater than 5.times.10.sup.-4 M, or greater than 1.times.10.sup.-3 M, as measured by biolayer interferometry.
4. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide.
5. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide binds to Protein A with a dissociation constant (K.sub.d) of less than 5.times.10' M, less than 1.times.10.sup.-6 M, less than 5.times.10.sup.-7 M, less than 1.times.10.sup.-7 M, less than 5.times.10.sup.-8 M, less than 1.times.10.sup.-8 M, less than 5.times.10.sup.-9 M, less than 1.times.10.sup.-9 M, less than 5.times.10.sup.-10 M, less than 1.times.10.sup.-10 M, less than 5.times.10.sup.-11 M, less than 1.times.10.sup.-11 M, less than 5.times.10.sup.-12 M, or less than 1.times.10.sup.-12 M, as measured by biolayer interferometry.
6. The polypeptide of any one of the preceding claims, wherein the companion animal species is canine, feline, or equine.
7. The polypeptide of any one of the preceding claims, wherein the wild-type IgG Fc polypeptide is a) a canine IgG-A Fc, IgG-B Fc, IgG-C Fc, or IgG-D Fc; b) an equine IgG1 Fc, IgG2 Fc, IgG3 Fc, IgG4 Fc, IgG5 Fc, IgG6 Fc, or IgG7 Fc; or c) a feline IgG1a Fc, IgG1b Fc, or IgG2 Fc.
8. A polypeptide comprising a variant IgG Fc polypeptide comprising at least one amino acid modification to a hinge region relative to a wild-type feline or equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide has increased recombinant production and/or increased hinge disulfide formation relative to the wild-type IgG Fc polypeptide, as determined by SDS-PAGE analysis under reducing and/or nonreducing conditions.
9. The polypeptide of any one the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118; b) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 3 of SEQ ID NO: 129; and/or c) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 20 of SEQ ID NO: 129.
10. The polypeptide of any one the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118; b) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at position 3 of SEQ ID NO: 129; and/or c) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at position 20 of SEQ ID NO: 129.
11. The polypeptide of any one the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises a proline at a position corresponding to position 16 or at position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118; b) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises a serine at a position corresponding to position 3 or at position 3 of SEQ ID NO: 129; and/or c) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises a proline at a position corresponding to position 20 or at position 20 of SEQ ID NO: 129.
12. The polypeptide of any one the preceding claims, wherein the variant IgG Fc polypeptide comprises a hinge region or a portion of a hinge region from an IgG Fc polypeptide of a different isotype.
13. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises a hinge region or a portion of a hinge region from a wild-type feline IgG-1a Fc polypeptide, from a wild-type feline IgG-1b Fc polypeptide, or from a wild-type equine IgG1 Fc polypeptide.
14. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises SEQ ID NO: 19, SEQ ID NO: 125 or SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID N: 134, SEQ ID NO: 135.
15. A polypeptide comprising an amino acid sequence of SEQ ID NO: 19, SEQ ID NO: 125 or SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID N: 134, SEQ ID NO: 135.
16. A polypeptide comprising a variant IgG2 Fc polypeptide comprising at least one amino acid substitution relative to a wild-type feline IgG2 Fc polypeptide, wherein the at least one amino acid substitution is a cysteine, and wherein the variant IgG2 Fc polypeptide is capable of forming at least one additional inter-chain disulfide linkage relative to the wild-type feline IgG2 Fc polypeptide.
17. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the at least one amino acid substitution is a cysteine, and wherein the variant IgG Fc polypeptide is capable of forming at least one additional inter-chain disulfide linkage relative to the wild-type feline IgG Fc polypeptide.
18. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises a cysteine at a position corresponding to position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, or position 16 of SEQ ID NO: 16.
19. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises a cysteine at a position corresponding to position 14 of SEQ ID NO: 16.
20. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises a cysteine at position 14 of SEQ ID NO: 16.
21. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide is at least 90% identical, at least 95% identical, at least 97% identical, or at least 99% identical to the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 100, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO:122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
22. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises SEQ ID NO: 17.
23. A polypeptide comprising an amino acid sequence of SEQ ID NO: 17.
24. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 205 of SEQ ID NO: 1, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 1; b) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 24 of SEQ ID NO: 3; c) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 4, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 4; d) an amino acid substitution at a position corresponding to position 15 of SEQ ID NO: 64, and/or an amino acid substitution at a position corresponding to position 203 of SEQ ID NO: 64; e) an amino acid substitution at a position corresponding to position 199 of SEQ ID NO: 67, and/or an amino acid substitution at a position corresponding to position 200 of SEQ ID NO: 67; and/or f) an amino acid substitution at a position corresponding to position 199 of SEQ ID NO: 68, an amino acid substitution at a position corresponding to position 200 of SEQ ID NO: 68, an amino acid substitution at a position corresponding to position 201 of SEQ ID NO: 68, and/or an amino acid substitution at a position corresponding to position 202 of SEQ ID NO: 68.
25. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an amino acid substitution at position 21 of SEQ ID NO: 1, an amino acid substitution at position 23 of SEQ ID NO: 1, an amino acid substitution at position 25 of SEQ ID NO: 1, an amino acid substitution at position 80 of SEQ ID NO: 1, an amino acid substitution at position 205 of SEQ ID NO: 1, and/or an amino acid substitution at position 207 of SEQ ID NO: 1; b) an amino acid substitution at position 21 of SEQ ID NO: 3, an amino acid substitution at position 23 of SEQ ID NO: 3, and/or an amino acid substitution at position 24 of SEQ ID NO: 3; c) an amino acid substitution at position 21 of SEQ ID NO: 4, an amino acid substitution at position 23 of SEQ ID NO: 4, an amino acid substitution at position 25 of SEQ ID NO: 4, an amino acid substitution at position 80 of SEQ ID NO: 4, and/or an amino acid substitution at position 207 of SEQ ID NO: 4; d) an amino acid substitution at position 15 of SEQ ID NO: 64, and/or an amino acid substitution at position 203 of SEQ ID NO: 64; e) an amino acid substitution at position 199 of SEQ ID NO: 67, and/or an amino acid substitution at position 200 of SEQ ID NO: 67; and/or f) an amino acid substitution at position 199 of SEQ ID NO: 68, an amino acid substitution at position 200 of SEQ ID NO: 68, an amino acid substitution at position 201 of SEQ ID NO: 68, and/or an amino acid substitution at position 202 of SEQ ID NO: 68.
26. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) a threonine at a position corresponding to position 21 of SEQ ID NO: 1, a leucine at a position corresponding to position 23 of SEQ ID NO: 1, an alanine at a position corresponding to position 25 of SEQ ID NO: 1, a glycine at a position corresponding to position 80 of SEQ ID NO: 1, an alanine at a position corresponding to position 205 of SEQ ID NO: 1, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 1; b) a threonine at a position corresponding to position 21 of SEQ ID NO: 3, a leucine at a position corresponding to position 23 of SEQ ID NO: 3, and/or an isoleucine at a position corresponding to position 24 of SEQ ID NO: 3; c) a threonine at a position corresponding to position 21 of SEQ ID NO: 4, a leucine at a position corresponding to position 23 of SEQ ID NO: 4, an alanine at a position corresponding to position 25 of SEQ ID NO: 4, a glycine at a position corresponding to position 80 of SEQ ID NO: 4, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 4; d) a threonine or a valine at a position corresponding to position 15 of SEQ ID NO: 64, and/or a tyrosine or a valine at a position corresponding to position 203 of SEQ ID NO: 64; e) a leucine at a position corresponding to position 199 of SEQ ID NO: 67, and/or a histidine at a position corresponding to position 200 of SEQ ID NO: 67; and/or f) a leucine at a position corresponding to position 199 of SEQ ID NO: 68, a histidine at a position corresponding to position 200 of SEQ ID NO: 68, an asparagine at a position corresponding to position 201 of SEQ ID NO: 68, and/or a histidine at a position corresponding to position 202 of SEQ ID NO: 68.
27. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) a threonine at position 21 of SEQ ID NO: 1, a leucine at position 23 of SEQ ID NO: 1, an alanine at position 25 of SEQ ID NO: 1, a glycine at position 80 of SEQ ID NO: 1, an alanine at position 205 of SEQ ID NO: 1, and/or a histidine at position 207 of SEQ ID NO: 1; b) a threonine at position 21 of SEQ ID NO: 3, a leucine at position 23 of SEQ ID NO: 3, and/or an isoleucine at position 24 of SEQ ID NO: 3; c) a threonine at a position 21 of SEQ ID NO: 4, a leucine at position 23 of SEQ ID NO: 4, an alanine at position 25 of SEQ ID NO: 4, a glycine at position 80 of SEQ ID NO: 4, and/or a histidine at position 207 of SEQ ID NO: 4; d) a threonine or a valine at position 15 of SEQ ID NO: 64, and/or a tyrosine or a valine at position 203 of SEQ ID NO: 64; e) a leucine at position 199 of SEQ ID NO: 67, and/or a histidine at position 200 of SEQ ID NO: 67; and/or f) a leucine at position 199 of SEQ ID NO: 68, a histidine at position 200 of SEQ ID NO: 68, an asparagine at position 201 of SEQ ID NO: 68, and/or a histidine at position 202 of SEQ ID NO: 68.
28. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of: a) SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, or SEQ ID NO: 84; or b) SEQ ID NO: 19, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 75, or SEQ ID NO: 76.
29. A polypeptide comprising an amino sequence of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 84, SEQ ID NO: 19, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 75, or SEQ ID NO: 76.
30. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 2, or an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 3; b) an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 63, an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 65, an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 66, or an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 69; or c) an amino acid substitution at a position corresponding to position 198 of SEQ ID NO: 80, or an amino acid substitution at a position corresponding to position 198 of SEQ ID NO: 81.
31. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an amino acid substitution at position 93 of SEQ ID NO: 2, or an amino acid substitution at position 93 of SEQ ID NO: 3; b) an amino acid substitution at position 87 of SEQ ID NO: 63, an amino acid substitution at position 87 of SEQ ID NO: 65, an amino acid substitution at position 87 of SEQ ID NO: 66, or an amino acid substitution at position 87 of SEQ ID NO: 69; or c) an amino acid substitution at position 198 of SEQ ID NO: 80, or an amino acid substitution at position 198 of SEQ ID NO: 81.
32. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an arginine at a position corresponding to position 93 of SEQ ID NO: 2, or an arginine at a position corresponding to position 93 of SEQ ID NO: 3; b) a serine at a position corresponding to position 87 of SEQ ID NO: 63, a serine substitution at a position corresponding to position 87 of SEQ ID NO: 65, a serine at a position corresponding to position 87 of SEQ ID NO: 66, or a serine at a position corresponding to position 87 of SEQ ID NO: 69; or c) an alanine at a position corresponding to position 198 of SEQ ID NO: 80, or an alanine at a position corresponding to position 198 of SEQ ID NO: 81.
33. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an arginine at position 93 of SEQ ID NO: 2, or an arginine at position 93 of SEQ ID NO: 3; b) a serine at position 87 of SEQ ID NO: 63, a serine at position 87 of SEQ ID NO: 65, a serine at position 87 of SEQ ID NO: 66, or a serine at position 87 of SEQ ID NO: 69; or c) an alanine at position 198 of SEQ ID NO: 80, or alanine at position 198 of SEQ ID NO: 81.
34. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises the amino acid sequence of: a) SEQ ID NO: 78, SEQ ID NO: 79, or SEQ ID NO: 84; or b) SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 74, or SEQ ID NO: 77; or c) SEQ ID NO: 82 or SEQ ID NO: 83.
35. A polypeptide comprising an amino sequence of SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 84, SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 77, SEQ ID NO: 82, or SEQ ID NO: 83.
36. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 2, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 2; or b) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 3.
37. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) an amino acid substitution at position 5 of SEQ ID NO: 2, an amino acid substitution at position 38 of SEQ ID NO: 2, an amino acid substitution at position 39 of SEQ ID NO: 2, an amino acid substitution at position 97 of SEQ ID NO: 2, and/or an amino acid substitution at position 98 of SEQ ID NO: 2; or b) an amino acid substitution at position 5 of SEQ ID NO: 3, an amino acid substitution at position 38 of SEQ ID NO: 3, an amino acid substitution at position 39 of SEQ ID NO: 3, an amino acid substitution at position 97 of SEQ ID NO: 3, and/or an amino acid substitution at position 98 of SEQ ID NO: 3.
38. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) a proline at a position corresponding to position 5 of SEQ ID NO: 2, a glycine at a position corresponding to position 38 of SEQ ID NO: 2, an arginine at a position corresponding to position 39 of SEQ ID NO: 2, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 2, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 2; or b) a proline at a position corresponding to position 5 of SEQ ID NO: 3, a glycine at a position corresponding to position 38 of SEQ ID NO: 3, an arginine at a position corresponding to position 39 of SEQ ID NO: 3, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 3, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 3.
39. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) a proline at position 5 of SEQ ID NO: 2, a glycine at position 38 of SEQ ID NO: 2, an arginine at position 39 of SEQ ID NO: 2, an isoleucine at position 97 of SEQ ID NO: 2, and/or a glycine at position 98 of SEQ ID NO: 2; or b) a proline at position 5 of SEQ ID NO: 3, a glycine at position 38 of SEQ ID NO: 3, an arginine at position 39 of SEQ ID NO: 3, an isoleucine at position 97 of SEQ ID NO: 3, and/or a glycine at position 98 of SEQ ID NO: 3.
40. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of: a) SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, or SEQ ID NO: 147; or b) SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
41. A polypeptide comprising an amino sequence of SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
42. A polypeptide comprising a variant IgG Fc polypeptide comprising: a) a tyrosine or a tryptophan at a position corresponding to position 138 of SEQ ID NO: 1, a tyrosine or a tryptophan at a position corresponding to position 137 of SEQ ID NO: 2, a tyrosine or a tryptophan at a position corresponding to position 137 of SEQ ID NO: 3, or a tyrosine or a tryptophan at a position corresponding to position 138 of SEQ ID NO: 4; or b) a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 16, a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 81 or SEQ ID NO: 118.
43. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) a tyrosine or a tryptophan at position 138 of SEQ ID NO: 1, a tyrosine or a tryptophan at position 137 of SEQ ID NO: 2, a tyrosine or a tryptophan at position 137 of SEQ ID NO: 3, or a tyrosine or a tryptophan at position 138 of SEQ ID NO: 4; or b) a tyrosine or a tryptophan at position 154 of SEQ ID NO: 16, a tyrosine or a tryptophan at position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 81 or SEQ ID NO: 118.
44. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
45. A polypeptide comprising an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
46. A contiguous polypeptide comprising the polypeptide of any one of the preceding claims and a glucagon-like peptide-1 (GLP1) polypeptide.
47. A contiguous polypeptide comprising the polypeptide of any one of the preceding claims and a glucagon polypeptide.
48. A polypeptide comprising a variant IgG Fc polypeptide comprising: a) a serine at a position corresponding to position 138 of SEQ ID NO: 1, a serine at a position corresponding to position 137 of SEQ ID NO: 2, a serine at a position corresponding to position 137 of SEQ ID NO: 3, a serine at a position corresponding to position 138 of SEQ ID NO: 4, a serine at a position corresponding to position 154 of SEQ ID NO: 16, a serine at a position corresponding to position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a serine at a position corresponding to position 154 of SEQ ID NO: 81 or SEQ ID NO: 118; b) an alanine at a position corresponding to position 140 of SEQ ID NO: 1, an alanine at a position corresponding to position 139 of SEQ ID NO: 2, an alanine at a position corresponding to position 139 of SEQ ID NO: 3, an alanine at a position corresponding to position 140 of SEQ ID NO: 4, an alanine at a position corresponding to position 156 of SEQ ID NO: 16, an alanine at a position corresponding to position 156 of SEQ ID NO: 80 or SEQ ID NO: 117, or an alanine at a position corresponding to position 156 of SEQ ID NO: 81 or SEQ ID NO: 118; and/or c) a threonine at a position corresponding to position 181 of SEQ ID NO: 1, a threonine at a position corresponding to position 180 of SEQ ID NO: 2, a threonine at a position corresponding to position 180 of SEQ ID NO: 3, a threonine at a position corresponding to position 181 of SEQ ID NO: 4, a threonine at a position corresponding to position 197 of SEQ ID NO: 16, a threonine at a position corresponding to position 197 of SEQ ID NO: 80 or SEQ ID NO: 117, or a threonine at a position corresponding to position 197 of SEQ ID NO: 81 or SEQ ID NO: 118.
49. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises: a) a serine at position 138 of SEQ ID NO: 1, a serine at position 137 of SEQ ID NO: 2, a serine at position 137 of SEQ ID NO: 3, a serine at position 138 of SEQ ID NO: 4, a serine at position 154 of SEQ ID NO: 16, a serine at position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a serine at position 154 of SEQ ID NO: 81 or SEQ ID NO: 118; b) an alanine at position 140 of SEQ ID NO: 1, an alanine at position 139 of SEQ ID NO: 2, an alanine at position 139 of SEQ ID NO: 3, an alanine at position 140 of SEQ ID NO: 4, an alanine at position 156 of SEQ ID NO: 16, an alanine at position 156 of SEQ ID NO: 80 or SEQ ID NO: 117, or an alanine at position 156 of SEQ ID NO: 81 or SEQ ID NO: 118; and/or; c) a threonine at position 181 of SEQ ID NO: 1, a threonine at position 181 of SEQ ID NO: 2, a threonine at position 181 of SEQ ID NO: 3, a threonine at position 181 of SEQ ID NO: 4, a threonine at position 197 of SEQ ID NO: 16, a threonine at position 197 of SEQ ID NO: 80 or SEQ ID NO: 117, or a threonine at position 197 of SEQ ID NO: 81 or SEQ ID NO: 118.
50. The polypeptide of any one of the preceding claims, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 116, SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 124.
51. A polypeptide comprising an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 116, SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 124.
52. The polypeptide of any one of the preceding claims, wherein the polypeptide is glycoslylated.
53. The polypeptide of any one of claims 1 to 51, wherein the polypeptide is aglycosylated.
54. A contiguous polypeptide comprising the polypeptide of any one of claims 48 to 53 and a glucagon-like peptide-1 (GLP1) polypeptide.
55. A contiguous polypeptide comprising the polypeptide of any one of claims 48 to 53 and a glucagon polypeptide.
56. A heterodimeric protein comprising the contiguous polypeptide of claim 46 and the contiguous polypeptide of claim 54.
57. A heterodimeric protein comprising the contiguous polypeptide of claim 47 and the contiguous polypeptide of claim 55.
58. The contiguous polypeptide or heterodimeric protein of any one of claims 46, 47, or 54 to 57, wherein the GLP1 polypeptide is a wild-type GLP1 polypeptide, optionally comprising the amino acid sequence of SEQ ID NO: 85.
59. The contiguous polypeptide or heterodimeric protein of any one of claims 46, 47, or 54 to 58, wherein the GLP1 polypeptide is a variant GLP1 polypeptide.
60. The contiguous polypeptide or heterodimeric protein of any one of claims 46, 47, or 54 to 59, wherein the GLP1 polypeptide comprises the amino acid sequence of SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 98, or SEQ ID NO: 99.
61. The contiguous polypeptide or heterodimeric protein of any one of claims 46, 47, or 54 to 60, wherein the glucagon polypeptide is a wild-type glucagon polypeptide, optionally comprising the amino acid sequence of SEQ ID NO: 21.
62. The contiguous polypeptide or heterodimeric protein of any one of claims 46, 47, or 54 to 61, wherein the glucagon polypeptide is a variant glucagon polypeptide.
63. A heterodimeric protein comprising: i) a first variant canine IgG Fc polypeptide comprising at least one amino acid modification relative to a first wild-type canine IgG Fc polypeptide and a second variant canine IgG Fc polypeptide comprising at least one amino acid modification relative to a second wild-type canine IgG Fc polypeptide; or ii) a first variant feline IgG Fc polypeptide comprising at least one amino acid modification relative to a first wild-type feline IgG Fc polypeptide and a second variant feline IgG Fc polypeptide comprising at least one amino acid modification relative to a second wild-type feline IgG Fc polypeptide, wherein: a) the first variant canine IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 138 of SEQ ID NO: 1, position 137 of SEQ ID NO: 2, position 137 of SEQ ID NO: 3, or position 138 of SEQ ID NO: 4; b) the second variant canine IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 138, position 140, and/or position 181 of SEQ ID NO: 1, position 137, position 139, and/or position 180 of SEQ ID NO: 2, position 137, position 139, and/or position 180 of SEQ ID NO: 3, or position 138, position 140, and/or position 181 of SEQ ID NO: 4; c) the first variant feline IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 154 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118; and/or d) the second variant feline IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 154, position 156, and/or position 197 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118.
64. The heterodimeric protein of claim 63, wherein the first wild-type canine IgG Fc polypeptide and the second wild-type canine IgG Fc polypeptide are from the same IgG subtype and/or the first wild-type feline IgG Fc polypeptide and the second wild-type feline IgG Fc polypeptide are from the same IgG subtype.
65. The heterodimeric protein of claim 63, wherein the first wild-type canine IgG Fc polypeptide and the second wild-type canine IgG Fc polypeptide are from a different IgG subtype and/or the first wild-type feline IgG Fc polypeptide and the second wild-type feline IgG Fc polypeptide are from the same IgG subtype.
66. The heterodimeric protein of any one of claims 63 to 65, wherein: a) the first variant canine IgG Fc polypeptide comprises a tyrosine or tryptophan at a position corresponding to position 138 of SEQ ID NO: 1, position 137 of SEQ ID NO: 2, position 137 of SEQ ID NO: 3, or position 138 of SEQ ID NO: 4; and/or b) the first variant feline IgG Fc polypeptide comprises a tryptophan at a position corresponding to position 154 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118.
67. The heterodimeric protein of any one of claims 63 to 66, wherein: a) the second variant canine IgG Fc polypeptide comprises a serine at a position corresponding to position 138, an alanine at a position corresponding to position 140, and/or a threonine at a position corresponding to position 181 of SEQ ID NO: 1, a serine at a position corresponding to position 137, an alanine at a position corresponding to position 139, and/or a threonine at a position corresponding to position 180 of SEQ ID NO: 2, a serine at a position corresponding to position 137, an alanine at a position corresponding to position 139, and/or a threonine at a position corresponding to position 180 of SEQ ID NO: 3, and/or a serine at a position corresponding to position 138, an alanine at a position corresponding to position 140, and/or a threonine at a position corresponding to position 181 of SEQ ID NO: 4; and/or b) the second variant feline IgG Fc polypeptide comprises a serine at a position corresponding to position 154, an alanine at a position corresponding to position 156, and/or a threonine at a position corresponding to position 197 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118.
68. The heterodimeric protein of any one of claims 63 to 67, wherein: a) the first variant canine IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, or SEQ ID NO: 115; and/or b) the first variant feline IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
69. The heterodimeric protein of any one of claims 63 to 68, wherein: a) the second variant canine IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, or SEQ ID NO: 116; and/or b) the second variant feline IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 123.
70. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 69, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises at least one additional amino acid modification relative to a wild-type IgG Fc polypeptide and has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide.
71. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 70, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 205 of SEQ ID NO: 1, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 1; b) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 24 of SEQ ID NO: 3; or c) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 4, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 4.
72. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 71, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) an amino acid substitution at position 21 of SEQ ID NO: 1, an amino acid substitution at position 23 of SEQ ID NO: 1, an amino acid substitution at position 25 of SEQ ID NO: 1, an amino acid substitution at position 80 of SEQ ID NO: 1, an amino acid substitution at position 205 of SEQ ID NO: 1, and/or an amino acid substitution at position 207 of SEQ ID NO: 1; b) an amino acid substitution at position 21 of SEQ ID NO: 3, an amino acid substitution at position 23 of SEQ ID NO: 3, and/or an amino acid substitution at position 24 of SEQ ID NO: 3; or c) an amino acid substitution at position 21 of SEQ ID NO: 4, an amino acid substitution at position 23 of SEQ ID NO: 4, an amino acid substitution at position 25 of SEQ ID NO: 4, an amino acid substitution at position 80 of SEQ ID NO: 4, and/or an amino acid substitution at position 207 of SEQ ID NO: 4.
73. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 72, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) a threonine at a position corresponding to position 21 of SEQ ID NO: 1, a leucine at a position corresponding to position 23 of SEQ ID NO: 1, an alanine at a position corresponding to position 25 of SEQ ID NO: 1, a glycine at a position corresponding to position 80 of SEQ ID NO: 1, an alanine at a position corresponding to position 205 of SEQ ID NO: 1, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 1; b) a threonine at a position corresponding to position 21 of SEQ ID NO: 3, a leucine at a position corresponding to position 23 of SEQ ID NO: 3, and/or an isoleucine at a position corresponding to position 24 of SEQ ID NO: 3; or c) a threonine at a position corresponding to position 21 of SEQ ID NO: 4, a leucine at a position corresponding to position 23 of SEQ ID NO: 4, an alanine at a position corresponding to position 25 of SEQ ID NO: 4, a glycine at a position corresponding to position 80 of SEQ ID NO: 4, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 4.
74. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 73, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) a threonine at position 21 of SEQ ID NO: 1, a leucine at position 23 of SEQ ID NO: 1, an alanine at position 25 of SEQ ID NO: 1, a glycine at position 80 of SEQ ID NO: 1, an alanine at position 205 of SEQ ID NO: 1, and/or a histidine at position 207 of SEQ ID NO: 1; b) a threonine at position 21 of SEQ ID NO: 3, a leucine at position 23 of SEQ ID NO: 3, and/or an isoleucine at position 24 of SEQ ID NO: 3; or c) a threonine at position 21 of SEQ ID NO: 4, a leucine at position 23 of SEQ ID NO: 4, an alanine at position 25 of SEQ ID NO: 4, a glycine at position 80 of SEQ ID NO: 4, and/or a histidine at position 207 of SEQ ID NO: 4.
75. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 74, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises at least one additional amino acid modification relative to a wild-type IgG Fc polypeptide and has decreased binding affinity to CD16 relative to the wild-type IgG Fc polypeptide.
76. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 75, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 2, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 2; or b) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 3.
77. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 76, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) an amino acid substitution at position 5 of SEQ ID NO: 2, an amino acid substitution at position 38 of SEQ ID NO: 2, an amino acid substitution at position 39 of SEQ ID NO: 2, an amino acid substitution at position 97 of SEQ ID NO: 2, and/or an amino acid substitution at position 98 of SEQ ID NO: 2; or b) an amino acid substitution at position 5 of SEQ ID NO: 3, an amino acid substitution at position 38 of SEQ ID NO: 3, an amino acid substitution at position 39 of SEQ ID NO: 3, an amino acid substitution at position 97 of SEQ ID NO: 3, and/or an amino acid substitution at position 98 of SEQ ID NO: 3.
78. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 77, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) a proline at a position corresponding to position 5 of SEQ ID NO: 2, a glycine at a position corresponding to position 38 of SEQ ID NO: 2, an arginine at a position corresponding to position 39 of SEQ ID NO: 2, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 2, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 2; or b) a proline at a position corresponding to position 5 of SEQ ID NO: 3, a glycine at a position corresponding to position 38 of SEQ ID NO: 3, an arginine at a position corresponding to position 39 of SEQ ID NO: 3, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 3, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 3.
79. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 78, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises: a) a proline at position 5 of SEQ ID NO: 2, a glycine at position 38 of SEQ ID NO: 2, an arginine at position 39 of SEQ ID NO: 2, an isoleucine at position 97 of SEQ ID NO: 2, and/or a glycine at position 98 of SEQ ID NO: 2; or b) a proline at position 5 of SEQ ID NO: 3, a glycine at position 38 of SEQ ID NO: 3, an arginine at position 39 of SEQ ID NO: 3, an isoleucine at position 97 of SEQ ID NO: 3, and/or a glycine at position 98 of SEQ ID NO: 3.
80. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 79, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises at least one additional amino acid modification relative to a wild-type canine IgG Fc polypeptide and has decreased binding affinity to C1q relative to the wild-type canine IgG Fc polypeptide.
81. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 80, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 2, or an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 3.
82. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 81, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an amino acid substitution at position 93 of SEQ ID NO: 2, or an amino acid substitution at position 93 of SEQ ID NO: 3.
83. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 82, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an arginine at a position corresponding to position 93 of SEQ ID NO: 2, or an arginine at a position corresponding to position 93 of SEQ ID NO: 3.
84. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 42 to 83, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an arginine at position 93 of SEQ ID NO: 2, or an arginine at position 93 of SEQ ID NO: 3.
85. The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 1 to 84, wherein the polypeptide is an antibody, an antibody fusion, or a fusion polypeptide.
86. A contiguous polypeptide comprising: a) a first glucagon-like peptide-1 (GLP1) polypeptide (GLP1A); b) a first linker (L1); c) an Fc polypeptide (Fc) of a companion animal species; d) optionally, a second linker (L2); and e) optionally, a second GLP1 polypeptide (GLP1B).
87. The contiguous polypeptide of claim 65 comprising: GLP1A-L1-Fc; or formula (I): Fc-L1-GLP1A. formula (II):
88. The contiguous polypeptide of claim 65 comprising: GLP1A-L1-Fc-L2-GLP1B. formula (III):
89. The contiguous polypeptide of any one of claims 86 to 88, wherein GLP1B, if present, comprises the same amino acid sequence as GLP1A.
90. A contiguous polypeptide comprising: a) a glucagon-like peptide-1 (GLP1) polypeptide; b) a first linker (L1); c) an Fc polypeptide (Fc); d) a second linker (L2); and e) a glucagon polypeptide (Gluc).
91. The contiguous polypeptide of claim 90 comprising: GLP1-L1-Fc-L2-Gluc; or Formula (IV): Gluc-L1-Fc-L2-GLP1. Formula (V):
92. The contiguous polypeptide of any one of claims 86 to 91, wherein GLP1A, GLP1, and/or GLP1B, if present, comprises a wild-type GLP1 polypeptide.
93. The contiguous polypeptide of any one of claims 86 to 92, wherein GLP1A, GLP1, and/or GLP1B, if present, comprises a variant GLP1 polypeptide.
94. The contiguous polypeptide of any one of claims 86 to 93, wherein GLP1A, GLP1, and/or GLP1B, if present, comprises an amino acid sequence of SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 98, or SEQ ID NO: 99.
95. The contiguous polypeptide of any one of claims 86 to 94, wherein the glucagon polypeptide comprises a wild-type glucagon polypeptide, optionally comprising the amino acid sequence of SEQ ID NO: 21.
96. The contiguous polypeptide of any one of claims 86 to 95, wherein the glucagon polypeptide is a variant glucagon polypeptide.
97. The contiguous polypeptide of any one of claims 86 to 96, wherein the Fc polypeptide is a human IgG Fc.
98. The contiguous polypeptide of any one of claims 86 to 97, wherein the Fc polypeptide is a human IgG1 Fc, IgG2 Fc, IgG3 Fc, or IgG4 Fc.
99. The contiguous polypeptide of any one of claims 86 to 98, wherein the Fc polypeptide is an Fc of a companion animal species.
100. The contiguous polypeptide of any one of claim 86 to 97 or 99, wherein the Fc polypeptide comprises: a) a canine IgG-A Fc, IgG-B Fc, IgG-C Fc, or IgG-D Fc; b) an equine IgG1 Fc, IgG2 Fc, IgG3 Fc, IgG4 Fc, IgG5 Fc, IgG6 Fc, or IgG7 Fc; or c) a feline IgG1a Fc, IgG1b Fc, or IgG2 Fc.
101. The contiguous polypeptide of any one of claims 86 to 100, wherein the Fc polypeptide is a wild-type IgG Fc polypeptide.
102. The contiguous polypeptide of any one of claims 86 to 100, wherein the Fc polypeptide is a variant IgG Fc polypeptide.
103. The contiguous polypeptide of any one of claims 86 to 102, wherein the Fc polypeptide comprises the polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of claims 1 to 84.
104. The contiguous polypeptide of any one of claims 85 to 102, wherein the contiguous polypeptide has a longer serum half-life than a wild-type GLP1 polypeptide.
105. The contiguous polypeptide of any one of claims 86 to 104, wherein L1 and L2, if present, each independently is a flexible linker.
106. The contiguous polypeptide of any one of claims 86 to 105, wherein the amino acid sequence of L1 and L2, if present, each independently comprises 100%, at least 95%, at least 90%, at least 85% serine and/or glycine amino acid residues.
107. The contiguous polypeptide of any one of claims 86 to 106, wherein the contiguous polypeptide comprises an extension at its C-terminus.
108. The contiguous polypeptide of any one of claims 86 to 107, wherein the contiguous polypeptide comprises a glycine residue, two glycine residues, three glycine residues, four glycine residues, five glycine residues, six glycine residues, seven glycine residues, eight glycine residues, or greater than eight glycine residues at its C-terminus.
109. The contiguous polypeptide of any one of claims 86 to 108, wherein the contiguous polypeptide comprises an amino acid sequence of SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, or SEQ ID NO: 95 at its C-terminus.
110. The contiguous polypeptide of any one of claims 86 to 109, wherein the contiguous polypeptide comprises: a) the amino acid sequence of SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; SEQ ID NO: 26; SEQ ID NO: 27; SEQ ID NO: 28; SEQ ID NO: 29; SEQ ID NO: 30; SEQ ID NO: 31; SEQ ID NO: 32; SEQ ID NO: 33; SEQ ID NO: 34; SEQ ID NO: 35; SEQ ID NO: 36; SEQ ID NO: 37; SEQ ID NO: 38; SEQ ID NO: 39; SEQ ID NO: 40; SEQ ID NO: 41; SEQ ID NO: 42; SEQ ID NO: 43; SEQ ID NO: 44; SEQ ID NO: 45; SEQ ID NO: 46; SEQ ID NO: 47, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, or SEQ ID NO: 106; or b) the amino acid sequence of SEQ ID NO: 52; SEQ ID NO: 53; SEQ ID NO: 54; SEQ ID NO: 55; SEQ ID NO: 56; SEQ ID NO: 57; SEQ ID NO: 58; or SEQ ID NO: 59.
111. A polypeptide comprising an amino acid sequence of SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; SEQ ID NO: 26; SEQ ID NO: 27; SEQ ID NO: 28; SEQ ID NO: 29; SEQ ID NO: 30; SEQ ID NO: 31; SEQ ID NO: 32; SEQ ID NO: 33; SEQ ID NO: 34; SEQ ID NO: 35; SEQ ID NO: 36; SEQ ID NO: 37; SEQ ID NO: 38; SEQ ID NO: 39; SEQ ID NO: 40; SEQ ID NO: 41; SEQ ID NO: 42; SEQ ID NO: 43; SEQ ID NO: 44; SEQ ID NO: 45; SEQ ID NO: 46; SEQ ID NO: 47, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 52; SEQ ID NO: 53; SEQ ID NO: 54; SEQ ID NO: 55; SEQ ID NO: 56; SEQ ID NO: 57; SEQ ID NO: 58; SEQ ID NO: 59; SEQ ID NO: 103; SEQ ID NO: 104, SEQ ID NO: 105, or SEQ ID NO: 106.
112. The polypeptide, the heterodimeric protein, or the contiguous polypeptide of any one of the preceding claims, wherein the at least one amino acid modification or substitution comprises an amino acid substitution with an amino acid derivative.
113. An isolated nucleic acid encoding the polypeptide, the heterodimeric protein, or the contiguous polypeptide of any one of the preceding claims.
114. A host cell comprising the nucleic acid of claim 113.
115. A method of producing a polypeptide comprising culturing the host cell of claim 114 and isolating the polypeptide.
116. A pharmaceutical composition comprising the polypeptide, the heterodimeric protein, or the contiguous polypeptide of any one of claims 1 to 112, and a pharmaceutically acceptable carrier.
117. A method of increasing production of cAMP in a cell, the method comprising exposing the cell to the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of claim 1 to 112 or 116 under conditions permissive for binding of the polypeptide, heterodimeric protein, or contiguous polypeptide to GLP1R.
118. The method of claim 117, wherein the cell is exposed to the polypeptide, heterodimeric protein, contiguous polypeptide, or the pharmaceutical composition ex vivo.
119. The method of claim 117, wherein the cell is exposed to the polypeptide, heterodimeric protein, contiguous polypeptide, or the pharmaceutical composition in vivo.
120. The method of any one of claims 118 to 119, wherein the cell is a human cell, a canine cell, a feline cell, or an equine cell.
121. A method of delivering a polypeptide to a subject comprising administering the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of claim 1 to 112 or 116 parenterally.
122. A method of delivering a polypeptide to a subject comprising administering the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of claim 1 to 112 or 116 by an intramuscular route, an intraperitoneal route, an intracerobrospinal route, a subcutaneous route, an intra-arterial route, an intrasynovial route, an intrathecal route, or an inhalation route.
123. A method of treating a subject having diabetes or obesity, the method comprising administering to the subject a therapeutically effective amount of the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of claim 1 to 112 or 116.
124. The method of claim 123, comprising administering insulin, a DPP4 inhibitor, a SGLT2 inhibitor, a biguanides sulfonylureas meglitinide derivative, an alpha-glucosidase inhibitor, a thiazolidinedion (TZD), an amylinomimetic, a bile acid sequestrant, a dopamine agonist.
125. The method of any one of claims 121 to 124, wherein the subject is a human subject.
126. The method of any one of claims 121 to 124, wherein the subject is a companion animal species.
127. The method of claim 126, wherein the companion animal species is canine, equine, or feline.
Description:
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application claims the benefit of priority of U.S. Provisional Application No. 62/545,858, filed Aug. 15, 2017, which is incorporated by reference herein in its entirety for any purpose.
FIELD
[0002] The present disclosure relates to variant IgG Fc polypeptides of companion animals with enhanced features, including increased Protein A binding (e.g., for ease of purification), decreased C1q binding (e.g., for reduced complement-mediated immune responses), decreased CD16 binding (e.g., for reduced antibody-dependent cellular cytotoxicity (ADCC) induction, increased stability, and/or the ability to form heterodimeric proteins. The variant IgG Fc polypeptides of the present disclosure may have broad applicability in companion animal therapeutics. For example, variant IgG Fc polypeptides may be used in the design and production of long-acting GLP1 polypeptides for treating, for example, diabetes, obesity, or related indications, in companion animals, such as canines, felines, and equines. In addition, variant IgG Fc polypeptides may be used in the design and production of antibodies or fusion proteins for treating various disorders in companion animals.
BACKGROUND
[0003] IgG Fc plays an important role in Fc-mediated functions though interactions with FcRn, Fc receptor, and C1q. In companion animals, various IgG subtypes possess differences in these functions, which are often considered when choosing a particular IgG antibody or IgG Fc fusion protein for therapeutic or diagnostic applications. For example, the ability of an IgG subtype to have weak or no measurable binding affinity to C1q or CD16 may be advantageous. In addition, IgG Fc's ability to bind Protein A may be useful for purification using a Protein A affinity purification platform.
[0004] However, most IgG Fc subtypes of canine, feline, and equine do not possess Protein A binding properties, weak or no measurable binding affinity to CD16, and weak or no measurable binding affinity to C1q. For example, of the four canine IgG Fc subtypes (IgG-A, IgG-B, IgG-C, and IgG-D), only canine IgG-B Fc has appreciable affinity to Protein A. Meanwhile only canine IgG-A Fc and IgG-D Fc have no or weak C1q binding or CD16 binding. Antibodies and Fc fusion proteins comprising variant IgG Fc polypeptides that have reduced binding to C1q and/or CD16, and/or that able to bind Protein A are desirable.
[0005] Glucagon-like peptide-1 (GLP1) is a potent antihyperglycemic hormone, which plays an important role in regulating blood glucose level. Native GLP1 has an in vivo half-life of approximately 2 minutes. Long-acting GLP1 polypeptides can be used to treat diabetes and obesity, prevent diabetes, control hyperglycemic conditions, lower lipids, treat conditions that would benefit from lowered blood glucose levels, suppress gastric or intestinal movement, slow gastric emptying, and/or decrease food intake. There remains a need for long-acting GLP1 polypeptides for treating high blood glucose or uncontrollable blood glucose-induced conditions in companion animals, such as canines, felines, and equines.
SUMMARY OF THE INVENTION
Embodiment 1
[0006] A polypeptide comprising a variant IgG Fc polypeptide comprising at least one amino acid modification relative to a wild-type IgG Fc polypeptide of a companion animal species, wherein the variant IgG Fc polypeptide has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide.
Embodiment 2
[0007] A polypeptide comprising a variant IgG Fc polypeptide comprising at least one amino acid modification relative to a wild-type IgG Fc polypeptide of a companion animal species, wherein the variant IgG Fc polypeptide has reduced binding affinity to C1q and/or CD16 relative to the wild-type IgG Fc polypeptide.
Embodiment 3
[0008] The polypeptide of embodiment 1 or embodiment 2, wherein the variant IgG Fc polypeptide binds to C1q and/or CD16 with a dissociation constant (K.sub.d) of greater than 5.times.10.sup.-6 M, greater than 1.times.10.sup.-5 M, greater than 5.times.10.sup.-5 M, greater than 1.times.10.sup.-4 M, greater than 5.times.10.sup.-4 M, or greater than 1.times.10.sup.-3 M, as measured by biolayer interferometry.
Embodiment 4
[0009] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide.
Embodiment 5
[0010] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide binds to Protein A with a dissociation constant (K.sub.d) of less than 5.times.10.sup.-6 M, less than 1.times.10.sup.-6 M, less than 5.times.10.sup.-7 M, less than 1.times.10.sup.-7 M, less than 5.times.10.sup.-8 M, less than 1.times.10.sup.-8 M, less than 5.times.10.sup.-9 M, less than 1.times.10.sup.-9 M, less than 5.times.10.sup.-10 M, less than 1.times.10.sup.-10 M, less than 5.times.10.sup.-11M, less than 1.times.10.sup.-11 M, less than 5.times.10.sup.-12 M, or less than 1.times.10.sup.-12 M, as measured by biolayer interferometry.
Embodiment 6
[0011] The polypeptide of any one of the preceding embodiments, wherein the companion animal species is canine, feline, or equine.
Embodiment 7
[0012] The polypeptide of any one of the preceding embodiments, wherein the wild-type IgG Fc polypeptide is
[0013] a) a canine IgG-A Fc, IgG-B Fc, IgG-C Fc, or IgG-D Fc;
[0014] b) an equine IgG1 Fc, IgG2 Fc, IgG3 Fc, IgG4 Fc, IgG5 Fc, IgG6 Fc, or IgG7 Fc; or
[0015] c) a feline IgG1a Fc, IgG1b Fc, or IgG2 Fc.
Embodiment 8
[0016] A polypeptide comprising a variant IgG Fc polypeptide comprising at least one amino acid modification to a hinge region relative to a wild-type feline or equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide has increased recombinant production and/or increased hinge disulfide formation relative to the wild-type IgG Fc polypeptide, as determined by SDS-PAGE analysis under reducing and/or nonreducing conditions.
Embodiment 9
[0017] The polypeptide of any one the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0018] a) at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118;
[0019] b) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 3 of SEQ ID NO: 129; and/or
[0020] c) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 20 of SEQ ID NO: 129.
Embodiment 10
[0021] The polypeptide of any one the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0022] a) at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118;
[0023] b) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at position 3 of SEQ ID NO: 129; and/or
[0024] c) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises an amino acid substitution at position 20 of SEQ ID NO: 129.
Embodiment 11
[0025] The polypeptide of any one the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0026] a) at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises a proline at a position corresponding to position 16 or at position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118;
[0027] b) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises a serine at a position corresponding to position 3 or at position 3 of SEQ ID NO: 129; and/or
[0028] c) at least one amino acid substitution relative to a wild-type equine IgG Fc polypeptide, wherein the variant IgG Fc polypeptide comprises a proline at a position corresponding to position 20 or at position 20 of SEQ ID NO: 129.
Embodiment 12
[0029] The polypeptide of any one the preceding embodiments, wherein the variant IgG Fc polypeptide comprises a hinge region or a portion of a hinge region from an IgG Fc polypeptide of a different isotype.
Embodiment 13
[0030] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises a hinge region or a portion of a hinge region from a wild-type feline IgG-1a Fc polypeptide, from a wild-type feline IgG-1b Fc polypeptide, or from a wild-type equine IgG1 Fc polypeptide.
Embodiment 14
[0031] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises SEQ ID NO: 19, SEQ ID NO: 125 or SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID N: 134, SEQ ID NO: 135.
Embodiment 15
[0032] A polypeptide comprising an amino acid sequence of SEQ ID NO: 19, SEQ ID NO: 125 or SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID N: 134, SEQ ID NO: 135.
Embodiment 16
[0033] A polypeptide comprising a variant IgG2 Fc polypeptide comprising at least one amino acid substitution relative to a wild-type feline IgG2 Fc polypeptide, wherein the at least one amino acid substitution is a cysteine, and wherein the variant IgG2 Fc polypeptide is capable of forming at least one additional inter-chain disulfide linkage relative to the wild-type feline IgG2 Fc polypeptide.
Embodiment 17
[0034] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises at least one amino acid substitution relative to a wild-type feline IgG Fc polypeptide, wherein the at least one amino acid substitution is a cysteine, and wherein the variant IgG Fc polypeptide is capable of forming at least one additional inter-chain disulfide linkage relative to the wild-type feline IgG Fc polypeptide.
Embodiment 18
[0035] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises a cysteine at a position corresponding to position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, or position 16 of SEQ ID NO: 16.
Embodiment 19
[0036] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises a cysteine at a position corresponding to position 14 of SEQ ID NO: 16.
Embodiment 20
[0037] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises a cysteine at position 14 of SEQ ID NO: 16.
Embodiment 21
[0038] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide is at least 90% identical, at least 95% identical, at least 97% identical, or at least 99% identical to the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 13, SEQ ID NO: 14, SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 70, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 75, SEQ ID NO: 76, SEQ ID NO: 77, SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 82, SEQ ID NO: 83, SEQ ID NO: 84, SEQ ID NO: 100, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 109, SEQ ID NO: 110, SEQ ID NO: 111, SEQ ID NO: 112, SEQ ID NO: 113, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 117, SEQ ID NO: 118, SEQ ID NO: 119, SEQ ID NO: 120, SEQ ID NO: 121, SEQ ID NO:122, SEQ ID NO: 123, SEQ ID NO: 124, SEQ ID NO: 125, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID NO: 134, SEQ ID NO: 135, SEQ ID NO: 136, SEQ ID NO: 137, SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 153, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
Embodiment 22
[0039] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises SEQ ID NO: 17.
Embodiment 23
[0040] A polypeptide comprising an amino acid sequence of SEQ ID NO: 17.
Embodiment 24
[0041] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0042] a) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 205 of SEQ ID NO: 1, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 1;
[0043] b) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 24 of SEQ ID NO: 3;
[0044] c) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 4, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 4;
[0045] d) an amino acid substitution at a position corresponding to position 15 of SEQ ID NO: 64, and/or an amino acid substitution at a position corresponding to position 203 of SEQ ID NO: 64;
[0046] e) an amino acid substitution at a position corresponding to position 199 of SEQ ID NO: 67, and/or an amino acid substitution at a position corresponding to position 200 of SEQ ID NO: 67; and/or
[0047] f) an amino acid substitution at a position corresponding to position 199 of SEQ ID NO: 68, an amino acid substitution at a position corresponding to position 200 of SEQ ID NO: 68, an amino acid substitution at a position corresponding to position 201 of SEQ ID NO: 68, and/or an amino acid substitution at a position corresponding to position 202 of SEQ ID NO: 68.
Embodiment 25
[0048] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0049] a) an amino acid substitution at position 21 of SEQ ID NO: 1, an amino acid substitution at position 23 of SEQ ID NO: 1, an amino acid substitution at position 25 of SEQ ID NO: 1, an amino acid substitution at position 80 of SEQ ID NO: 1, an amino acid substitution at position 205 of SEQ ID NO: 1, and/or an amino acid substitution at position 207 of SEQ ID NO: 1;
[0050] b) an amino acid substitution at position 21 of SEQ ID NO: 3, an amino acid substitution at position 23 of SEQ ID NO: 3, and/or an amino acid substitution at position 24 of SEQ ID NO: 3;
[0051] c) an amino acid substitution at position 21 of SEQ ID NO: 4, an amino acid substitution at position 23 of SEQ ID NO: 4, an amino acid substitution at position 25 of SEQ ID NO: 4, an amino acid substitution at position 80 of SEQ ID NO: 4, and/or an amino acid substitution at position 207 of SEQ ID NO: 4;
[0052] d) an amino acid substitution at position 15 of SEQ ID NO: 64, and/or an amino acid substitution at position 203 of SEQ ID NO: 64;
[0053] e) an amino acid substitution at position 199 of SEQ ID NO: 67, and/or an amino acid substitution at position 200 of SEQ ID NO: 67; and/or
[0054] f) an amino acid substitution at position 199 of SEQ ID NO: 68, an amino acid substitution at position 200 of SEQ ID NO: 68, an amino acid substitution at position 201 of SEQ ID NO: 68, and/or an amino acid substitution at position 202 of SEQ ID NO: 68.
Embodiment 26
[0055] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0056] a) a threonine at a position corresponding to position 21 of SEQ ID NO: 1, a leucine at a position corresponding to position 23 of SEQ ID NO: 1, an alanine at a position corresponding to position 25 of SEQ ID NO: 1, a glycine at a position corresponding to position 80 of SEQ ID NO: 1, an alanine at a position corresponding to position 205 of SEQ ID NO: 1, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 1;
[0057] b) a threonine at a position corresponding to position 21 of SEQ ID NO: 3, a leucine at a position corresponding to position 23 of SEQ ID NO: 3, and/or an isoleucine at a position corresponding to position 24 of SEQ ID NO: 3;
[0058] c) a threonine at a position corresponding to position 21 of SEQ ID NO: 4, a leucine at a position corresponding to position 23 of SEQ ID NO: 4, an alanine at a position corresponding to position 25 of SEQ ID NO: 4, a glycine at a position corresponding to position 80 of SEQ ID NO: 4, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 4;
[0059] d) a threonine or a valine at a position corresponding to position 15 of SEQ ID NO: 64, and/or a tyrosine or a valine at a position corresponding to position 203 of SEQ ID NO: 64;
[0060] e) a leucine at a position corresponding to position 199 of SEQ ID NO: 67, and/or a histidine at a position corresponding to position 200 of SEQ ID NO: 67; and/or
[0061] f) a leucine at a position corresponding to position 199 of SEQ ID NO: 68, a histidine at a position corresponding to position 200 of SEQ ID NO: 68, an asparagine at a position corresponding to position 201 of SEQ ID NO: 68, and/or a histidine at a position corresponding to position 202 of SEQ ID NO: 68.
Embodiment 27
[0062] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0063] a) a threonine at position 21 of SEQ ID NO: 1, a leucine at position 23 of SEQ ID NO: 1, an alanine at position 25 of SEQ ID NO: 1, a glycine at position 80 of SEQ ID NO: 1, an alanine at position 205 of SEQ ID NO: 1, and/or a histidine at position 207 of SEQ ID NO: 1;
[0064] b) a threonine at position 21 of SEQ ID NO: 3, a leucine at position 23 of SEQ ID NO: 3, and/or an isoleucine at position 24 of SEQ ID NO: 3;
[0065] c) a threonine at a position 21 of SEQ ID NO: 4, a leucine at position 23 of SEQ ID NO: 4, an alanine at position 25 of SEQ ID NO: 4, a glycine at position 80 of SEQ ID NO: 4, and/or a histidine at position 207 of SEQ ID NO: 4;
[0066] d) a threonine or a valine at position 15 of SEQ ID NO: 64, and/or a tyrosine or a valine at position 203 of SEQ ID NO: 64;
[0067] e) a leucine at position 199 of SEQ ID NO: 67, and/or a histidine at position 200 of SEQ ID NO: 67; and/or
[0068] f) a leucine at position 199 of SEQ ID NO: 68, a histidine at position 200 of SEQ ID NO: 68, an asparagine at position 201 of SEQ ID NO: 68, and/or a histidine at position 202 of SEQ ID NO: 68.
Embodiment 28
[0069] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of:
[0070] a) SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, or SEQ ID NO: 84; or
[0071] b) SEQ ID NO: 19, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 75, or SEQ ID NO: 76.
Embodiment 29
[0072] A polypeptide comprising an amino sequence of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, SEQ ID NO: 84, SEQ ID NO: 19, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 75, or SEQ ID NO: 76.
Embodiment 30
[0073] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0074] a) an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 2, or an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 3;
[0075] b) an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 63, an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 65, an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 66, or an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 69; or
[0076] c) an amino acid substitution at a position corresponding to position 198 of SEQ ID NO: 80, or an amino acid substitution at a position corresponding to position 198 of SEQ ID NO: 81.
Embodiment 31
[0077] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0078] a) an amino acid substitution at position 93 of SEQ ID NO: 2, or an amino acid substitution at position 93 of SEQ ID NO: 3;
[0079] b) an amino acid substitution at position 87 of SEQ ID NO: 63, an amino acid substitution at position 87 of SEQ ID NO: 65, an amino acid substitution at position 87 of SEQ ID NO: 66, or an amino acid substitution at position 87 of SEQ ID NO: 69; or
[0080] c) an amino acid substitution at position 198 of SEQ ID NO: 80, or an amino acid substitution at position 198 of SEQ ID NO: 81.
Embodiment 32
[0081] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0082] a) an arginine at a position corresponding to position 93 of SEQ ID NO: 2, or an arginine at a position corresponding to position 93 of SEQ ID NO: 3;
[0083] b) a serine at a position corresponding to position 87 of SEQ ID NO: 63, a serine substitution at a position corresponding to position 87 of SEQ ID NO: 65, a serine at a position corresponding to position 87 of SEQ ID NO: 66, or a serine at a position corresponding to position 87 of SEQ ID NO: 69; or
[0084] c) an alanine at a position corresponding to position 198 of SEQ ID NO: 80, or an alanine at a position corresponding to position 198 of SEQ ID NO: 81.
Embodiment 33
[0085] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0086] a) an arginine at position 93 of SEQ ID NO: 2, or an arginine at position 93 of SEQ ID NO: 3;
[0087] b) a serine at position 87 of SEQ ID NO: 63, a serine at position 87 of SEQ ID NO: 65, a serine at position 87 of SEQ ID NO: 66, or a serine at position 87 of SEQ ID NO: 69; or
[0088] c) an alanine at position 198 of SEQ ID NO: 80, or alanine at position 198 of SEQ ID NO: 81.
Embodiment 34
[0089] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises the amino acid sequence of:
[0090] a) SEQ ID NO: 78, SEQ ID NO: 79, or SEQ ID NO: 84; or
[0091] b) SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 74, or SEQ ID NO: 77; or
[0092] c) SEQ ID NO: 82 or SEQ ID NO: 83.
Embodiment 35
[0093] A polypeptide comprising an amino sequence of SEQ ID NO: 78, SEQ ID NO: 79, SEQ ID NO: 84, SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 74, SEQ ID NO: 77, SEQ ID NO: 82, or SEQ ID NO: 83.
Embodiment 36
[0094] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0095] a) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 2, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 2; or
[0096] b) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 3.
Embodiment 37
[0097] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0098] a) an amino acid substitution at position 5 of SEQ ID NO: 2, an amino acid substitution at position 38 of SEQ ID NO: 2, an amino acid substitution at position 39 of SEQ ID NO: 2, an amino acid substitution at position 97 of SEQ ID NO: 2, and/or an amino acid substitution at position 98 of SEQ ID NO: 2; or
[0099] b) an amino acid substitution at position 5 of SEQ ID NO: 3, an amino acid substitution at position 38 of SEQ ID NO: 3, an amino acid substitution at position 39 of SEQ ID NO: 3, an amino acid substitution at position 97 of SEQ ID NO: 3, and/or an amino acid substitution at position 98 of SEQ ID NO: 3.
Embodiment 38
[0100] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0101] a) a proline at a position corresponding to position 5 of SEQ ID NO: 2, a glycine at a position corresponding to position 38 of SEQ ID NO: 2, an arginine at a position corresponding to position 39 of SEQ ID NO: 2, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 2, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 2; or
[0102] b) a proline at a position corresponding to position 5 of SEQ ID NO: 3, a glycine at a position corresponding to position 38 of SEQ ID NO: 3, an arginine at a position corresponding to position 39 of SEQ ID NO: 3, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 3, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 3.
Embodiment 39
[0103] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0104] a) a proline at position 5 of SEQ ID NO: 2, a glycine at position 38 of SEQ ID NO: 2, an arginine at position 39 of SEQ ID NO: 2, an isoleucine at position 97 of SEQ ID NO: 2, and/or a glycine at position 98 of SEQ ID NO: 2; or
[0105] b) a proline at position 5 of SEQ ID NO: 3, a glycine at position 38 of SEQ ID NO: 3, an arginine at position 39 of SEQ ID NO: 3, an isoleucine at position 97 of SEQ ID NO: 3, and/or a glycine at position 98 of SEQ ID NO: 3.
Embodiment 40
[0106] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of:
[0107] a) SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, or SEQ ID NO: 147; or
[0108] b) SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
Embodiment 41
[0109] A polypeptide comprising an amino sequence of SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID NO: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
Embodiment 42
[0110] A polypeptide comprising a variant IgG Fc polypeptide comprising:
[0111] a) a tyrosine or a tryptophan at a position corresponding to position 138 of SEQ ID NO: 1, a tyrosine or a tryptophan at a position corresponding to position 137 of SEQ ID NO: 2, a tyrosine or a tryptophan at a position corresponding to position 137 of SEQ ID NO: 3, or a tyrosine or a tryptophan at a position corresponding to position 138 of SEQ ID NO: 4; or
[0112] b) a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 16, a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 81 or SEQ ID NO: 118.
Embodiment 43
[0113] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0114] a) a tyrosine or a tryptophan at position 138 of SEQ ID NO: 1, a tyrosine or a tryptophan at position 137 of SEQ ID NO: 2, a tyrosine or a tryptophan at position 137 of SEQ ID NO: 3, or a tyrosine or a tryptophan at position 138 of SEQ ID NO: 4; or
[0115] b) a tyrosine or a tryptophan at position 154 of SEQ ID NO: 16, a tyrosine or a tryptophan at position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a tyrosine or a tryptophan at a position corresponding to position 154 of SEQ ID NO: 81 or SEQ ID NO: 118.
Embodiment 44
[0116] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
Embodiment 45
[0117] A polypeptide comprising an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
Embodiment 46
[0118] A contiguous polypeptide comprising the polypeptide of any one of the preceding embodiments and a glucagon-like peptide-1 (GLP1) polypeptide.
Embodiment 47
[0119] A contiguous polypeptide comprising the polypeptide of any one of the preceding embodiments and a glucagon polypeptide.
Embodiment 48
[0120] A polypeptide comprising a variant IgG Fc polypeptide comprising:
[0121] a) a serine at a position corresponding to position 138 of SEQ ID NO: 1, a serine at a position corresponding to position 137 of SEQ ID NO: 2, a serine at a position corresponding to position 137 of SEQ ID NO: 3, a serine at a position corresponding to position 138 of SEQ ID NO: 4, a serine at a position corresponding to position 154 of SEQ ID NO: 16, a serine at a position corresponding to position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a serine at a position corresponding to position 154 of SEQ ID NO: 81 or SEQ ID NO: 118;
[0122] b) an alanine at a position corresponding to position 140 of SEQ ID NO: 1, an alanine at a position corresponding to position 139 of SEQ ID NO: 2, an alanine at a position corresponding to position 139 of SEQ ID NO: 3, an alanine at a position corresponding to position 140 of SEQ ID NO: 4, an alanine at a position corresponding to position 156 of SEQ ID NO: 16, an alanine at a position corresponding to position 156 of SEQ ID NO: 80 or SEQ ID NO: 117, or an alanine at a position corresponding to position 156 of SEQ ID NO: 81 or SEQ ID NO: 118; and/or
[0123] c) a threonine at a position corresponding to position 181 of SEQ ID NO: 1, a threonine at a position corresponding to position 180 of SEQ ID NO: 2, a threonine at a position corresponding to position 180 of SEQ ID NO: 3, a threonine at a position corresponding to position 181 of SEQ ID NO: 4, a threonine at a position corresponding to position 197 of SEQ ID NO: 16, a threonine at a position corresponding to position 197 of SEQ ID NO: 80 or SEQ ID NO: 117, or a threonine at a position corresponding to position 197 of SEQ ID NO: 81 or SEQ ID NO: 118.
Embodiment 49
[0124] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises:
[0125] a) a serine at position 138 of SEQ ID NO: 1, a serine at position 137 of SEQ ID NO: 2, a serine at position 137 of SEQ ID NO: 3, a serine at position 138 of SEQ ID NO: 4, a serine at position 154 of SEQ ID NO: 16, a serine at position 154 of SEQ ID NO: 80 or SEQ ID NO: 117, or a serine at position 154 of SEQ ID NO: 81 or SEQ ID NO: 118;
[0126] b) an alanine at position 140 of SEQ ID NO: 1, an alanine at position 139 of SEQ ID NO: 2, an alanine at position 139 of SEQ ID NO: 3, an alanine at position 140 of SEQ ID NO: 4, an alanine at position 156 of SEQ ID NO: 16, an alanine at position 156 of SEQ ID NO: 80 or SEQ ID NO: 117, or an alanine at position 156 of SEQ ID NO: 81 or SEQ ID NO: 118; and/or;
[0127] c) a threonine at position 181 of SEQ ID NO: 1, a threonine at position 181 of SEQ ID NO: 2, a threonine at position 181 of SEQ ID NO: 3, a threonine at position 181 of SEQ ID NO: 4, a threonine at position 197 of SEQ ID NO: 16, a threonine at position 197 of SEQ ID NO: 80 or SEQ ID NO: 117, or a threonine at position 197 of SEQ ID NO: 81 or SEQ ID NO: 118.
Embodiment 50
[0128] The polypeptide of any one of the preceding embodiments, wherein the variant IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 116, SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 124.
Embodiment 51
[0129] A polypeptide comprising an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 116, SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 124.
Embodiment 52
[0130] The polypeptide of any one of the preceding embodiments, wherein the polypeptide is glycoslylated.
Embodiment 53
[0131] The polypeptide of any one of embodiments 1 to 51, wherein the polypeptide is a glycosylated.
Embodiment 54
[0132] A contiguous polypeptide comprising the polypeptide of any one of embodiments 48 to 53 and a glucagon-like peptide-1 (GLP1) polypeptide.
Embodiment 55
[0133] A contiguous polypeptide comprising the polypeptide of any one of embodiments 48 to 53 and a glucagon polypeptide.
Embodiment 56
[0134] A heterodimeric protein comprising the contiguous polypeptide of embodiment 46 and the contiguous polypeptide of embodiment 54.
Embodiment 57
[0135] A heterodimeric protein comprising the contiguous polypeptide of embodiment 47 and the contiguous polypeptide of embodiment 55.
Embodiment 58
[0136] The contiguous polypeptide or heterodimeric protein of any one of embodiments 46, 47, or 54 to 57, wherein the GLP1 polypeptide is a wild-type GLP1 polypeptide, optionally comprising the amino acid sequence of SEQ ID NO: 85.
Embodiment 59
[0137] The contiguous polypeptide or heterodimeric protein of any one of embodiments 46, 47, or 54 to 58, wherein the GLP1 polypeptide is a variant GLP1 polypeptide.
Embodiment 60
[0138] The contiguous polypeptide or heterodimeric protein of any one of embodiments 46, 47, or 54 to 59, wherein the GLP1 polypeptide comprises the amino acid sequence of SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 98, or SEQ ID NO: 99.
Embodiment 61
[0139] The contiguous polypeptide or heterodimeric protein of any one of embodiments 46, 47, or 54 to 60, wherein the glucagon polypeptide is a wild-type glucagon polypeptide, optionally comprising the amino acid sequence of SEQ ID NO: 21.
Embodiment 62
[0140] The contiguous polypeptide or heterodimeric protein of any one of embodiments 46, 47, or 54 to 61, wherein the glucagon polypeptide is a variant glucagon polypeptide.
Embodiment 63
[0141] A heterodimeric protein comprising:
i) a first variant canine IgG Fc polypeptide comprising at least one amino acid modification relative to a first wild-type canine IgG Fc polypeptide and a second variant canine IgG Fc polypeptide comprising at least one amino acid modification relative to a second wild-type canine IgG Fc polypeptide; or ii) a first variant feline IgG Fc polypeptide comprising at least one amino acid modification relative to a first wild-type feline IgG Fc polypeptide and a second variant feline IgG Fc polypeptide comprising at least one amino acid modification relative to a second wild-type feline IgG Fc polypeptide, wherein:
[0142] a) the first variant canine IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 138 of SEQ ID NO: 1, position 137 of SEQ ID NO: 2, position 137 of SEQ ID NO: 3, or position 138 of SEQ ID NO: 4;
[0143] b) the second variant canine IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 138, position 140, and/or position 181 of SEQ ID NO: 1, position 137, position 139, and/or position 180 of SEQ ID NO: 2, position 137, position 139, and/or position 180 of SEQ ID NO: 3, or position 138, position 140, and/or position 181 of SEQ ID NO: 4;
[0144] c) the first variant feline IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 154 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118; and/or
[0145] d) the second variant feline IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 154, position 156, and/or position 197 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118.
Embodiment 64
[0146] The heterodimeric protein of embodiment 63, wherein the first wild-type canine IgG Fc polypeptide and the second wild-type canine IgG Fc polypeptide are from the same IgG subtype and/or the first wild-type feline IgG Fc polypeptide and the second wild-type feline IgG Fc polypeptide are from the same IgG subtype.
Embodiment 65
[0147] The heterodimeric protein of embodiment 63, wherein the first wild-type canine IgG Fc polypeptide and the second wild-type canine IgG Fc polypeptide are from a different IgG subtype and/or the first wild-type feline IgG Fc polypeptide and the second wild-type feline IgG Fc polypeptide are from the same IgG subtype.
Embodiment 66
[0148] The heterodimeric protein of any one of embodiments 63 to 65, wherein:
[0149] a) the first variant canine IgG Fc polypeptide comprises a tyrosine or tryptophan at a position corresponding to position 138 of SEQ ID NO: 1, position 137 of SEQ ID NO: 2, position 137 of SEQ ID NO: 3, or position 138 of SEQ ID NO: 4; and/or
[0150] b) the first variant feline IgG Fc polypeptide comprises a tryptophan at a position corresponding to position 154 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118.
Embodiment 67
[0151] The heterodimeric protein of any one of embodiments 63 to 66, wherein:
[0152] a) the second variant canine IgG Fc polypeptide comprises a serine at a position corresponding to position 138, an alanine at a position corresponding to position 140, and/or a threonine at a position corresponding to position 181 of SEQ ID NO: 1, a serine at a position corresponding to position 137, an alanine at a position corresponding to position 139, and/or a threonine at a position corresponding to position 180 of SEQ ID NO: 2, a serine at a position corresponding to position 137, an alanine at a position corresponding to position 139, and/or a threonine at a position corresponding to position 180 of SEQ ID NO: 3, and/or a serine at a position corresponding to position 138, an alanine at a position corresponding to position 140, and/or a threonine at a position corresponding to position 181 of SEQ ID NO: 4; and/or
[0153] b) the second variant feline IgG Fc polypeptide comprises a serine at a position corresponding to position 154, an alanine at a position corresponding to position 156, and/or a threonine at a position corresponding to position 197 of SEQ ID NO: 6, of SEQ ID NO: 80, of SEQ ID NO: 81, of SEQ ID NO: 117, or of SEQ ID NO: 118.
Embodiment 68
[0154] The heterodimeric protein of any one of embodiments 63 to 67, wherein:
[0155] a) the first variant canine IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, or SEQ ID NO: 115; and/or
[0156] b) the first variant feline IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
Embodiment 69
[0157] The heterodimeric protein of any one of embodiments 63 to 68, wherein:
[0158] a) the second variant canine IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, or SEQ ID NO: 116; and/or
[0159] b) the second variant feline IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 123.
Embodiment 70
[0160] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 69, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises at least one additional amino acid modification relative to a wild-type IgG Fc polypeptide and has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide.
Embodiment 71
[0161] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 70, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0162] a) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 1, an amino acid substitution at a position corresponding to position 205 of SEQ ID NO: 1, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 1;
[0163] b) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 24 of SEQ ID NO: 3; or
[0164] c) an amino acid substitution at a position corresponding to position 21 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 23 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 25 of SEQ ID NO: 4, an amino acid substitution at a position corresponding to position 80 of SEQ ID NO: 4, and/or an amino acid substitution at a position corresponding to position 207 of SEQ ID NO: 4.
Embodiment 72
[0165] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 71, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0166] a) an amino acid substitution at position 21 of SEQ ID NO: 1, an amino acid substitution at position 23 of SEQ ID NO: 1, an amino acid substitution at position 25 of SEQ ID NO: 1, an amino acid substitution at position 80 of SEQ ID NO: 1, an amino acid substitution at position 205 of SEQ ID NO: 1, and/or an amino acid substitution at position 207 of SEQ ID NO: 1;
[0167] b) an amino acid substitution at position 21 of SEQ ID NO: 3, an amino acid substitution at position 23 of SEQ ID NO: 3, and/or an amino acid substitution at position 24 of SEQ ID NO: 3; or
[0168] c) an amino acid substitution at position 21 of SEQ ID NO: 4, an amino acid substitution at position 23 of SEQ ID NO: 4, an amino acid substitution at position 25 of SEQ ID NO: 4, an amino acid substitution at position 80 of SEQ ID NO: 4, and/or an amino acid substitution at position 207 of SEQ ID NO: 4.
Embodiment 73
[0169] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 72, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0170] a) a threonine at a position corresponding to position 21 of SEQ ID NO: 1, a leucine at a position corresponding to position 23 of SEQ ID NO: 1, an alanine at a position corresponding to position 25 of SEQ ID NO: 1, a glycine at a position corresponding to position 80 of SEQ ID NO: 1, an alanine at a position corresponding to position 205 of SEQ ID NO: 1, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 1;
[0171] b) a threonine at a position corresponding to position 21 of SEQ ID NO: 3, a leucine at a position corresponding to position 23 of SEQ ID NO: 3, and/or an isoleucine at a position corresponding to position 24 of SEQ ID NO: 3; or
[0172] c) a threonine at a position corresponding to position 21 of SEQ ID NO: 4, a leucine at a position corresponding to position 23 of SEQ ID NO: 4, an alanine at a position corresponding to position 25 of SEQ ID NO: 4, a glycine at a position corresponding to position 80 of SEQ ID NO: 4, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 4.
Embodiment 74
[0173] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 73, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0174] a) a threonine at position 21 of SEQ ID NO: 1, a leucine at position 23 of SEQ ID NO: 1, an alanine at position 25 of SEQ ID NO: 1, a glycine at position 80 of SEQ ID NO: 1, an alanine at position 205 of SEQ ID NO: 1, and/or a histidine at position 207 of SEQ ID NO: 1;
[0175] b) a threonine at position 21 of SEQ ID NO: 3, a leucine at position 23 of SEQ ID NO: 3, and/or an isoleucine at position 24 of SEQ ID NO: 3; or
[0176] c) a threonine at position 21 of SEQ ID NO: 4, a leucine at position 23 of SEQ ID NO: 4, an alanine at position 25 of SEQ ID NO: 4, a glycine at position 80 of SEQ ID NO: 4, and/or a histidine at position 207 of SEQ ID NO: 4.
Embodiment 75
[0177] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 74, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises at least one additional amino acid modification relative to a wild-type IgG Fc polypeptide and has decreased binding affinity to CD16 relative to the wild-type IgG Fc polypeptide.
Embodiment 76
[0178] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 75, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0179] a) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 2, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 2, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 2; or
[0180] b) an amino acid substitution at a position corresponding to position 5 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 38 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 39 of SEQ ID NO: 3, an amino acid substitution at a position corresponding to position 97 of SEQ ID NO: 3, and/or an amino acid substitution at a position corresponding to position 98 of SEQ ID NO: 3.
Embodiment 77
[0181] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 76, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0182] a) an amino acid substitution at position 5 of SEQ ID NO: 2, an amino acid substitution at position 38 of SEQ ID NO: 2, an amino acid substitution at position 39 of SEQ ID NO: 2, an amino acid substitution at position 97 of SEQ ID NO: 2, and/or an amino acid substitution at position 98 of SEQ ID NO: 2; or
[0183] b) an amino acid substitution at position 5 of SEQ ID NO: 3, an amino acid substitution at position 38 of SEQ ID NO: 3, an amino acid substitution at position 39 of SEQ ID NO: 3, an amino acid substitution at position 97 of SEQ ID NO: 3, and/or an amino acid substitution at position 98 of SEQ ID NO: 3.
Embodiment 78
[0184] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 77, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0185] a) a proline at a position corresponding to position 5 of SEQ ID NO: 2, a glycine at a position corresponding to position 38 of SEQ ID NO: 2, an arginine at a position corresponding to position 39 of SEQ ID NO: 2, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 2, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 2; or
[0186] b) a proline at a position corresponding to position 5 of SEQ ID NO: 3, a glycine at a position corresponding to position 38 of SEQ ID NO: 3, an arginine at a position corresponding to position 39 of SEQ ID NO: 3, an isoleucine at a position corresponding to position 97 of SEQ ID NO: 3, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 3.
Embodiment 79
[0187] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 78, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises:
[0188] a) a proline at position 5 of SEQ ID NO: 2, a glycine at position 38 of SEQ ID NO: 2, an arginine at position 39 of SEQ ID NO: 2, an isoleucine at position 97 of SEQ ID NO: 2, and/or a glycine at position 98 of SEQ ID NO: 2; or
[0189] b) a proline at position 5 of SEQ ID NO: 3, a glycine at position 38 of SEQ ID NO: 3, an arginine at position 39 of SEQ ID NO: 3, an isoleucine at position 97 of SEQ ID NO: 3, and/or a glycine at position 98 of SEQ ID NO: 3.
Embodiment 80
[0190] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 79, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises at least one additional amino acid modification relative to a wild-type canine IgG Fc polypeptide and has decreased binding affinity to C1q relative to the wild-type canine IgG Fc polypeptide.
Embodiment 81
[0191] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 80, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 2, or an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 3.
Embodiment 82
[0192] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 81, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an amino acid substitution at position 93 of SEQ ID NO: 2, or an amino acid substitution at position 93 of SEQ ID NO: 3.
Embodiment 83
[0193] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 82, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an arginine at a position corresponding to position 93 of SEQ ID NO: 2, or an arginine at a position corresponding to position 93 of SEQ ID NO: 3.
Embodiment 84
[0194] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 42 to 83, wherein the variant IgG Fc polypeptide, the first variant IgG Fc polypeptide, and/or the second variant IgG Fc polypeptide comprises an arginine at position 93 of SEQ ID NO: 2, or an arginine at position 93 of SEQ ID NO: 3.
Embodiment 85
[0195] The polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 1 to 84, wherein the polypeptide is an antibody, an antibody fusion, or a fusion polypeptide.
Embodiment 86
[0196] A contiguous polypeptide comprising:
[0197] a) a first glucagon-like peptide-1 (GLP1) polypeptide (GLP1A);
[0198] b) a first linker (L1);
[0199] c) an Fc polypeptide (Fc) of a companion animal species;
[0200] d) optionally, a second linker (L2); and
[0201] e) optionally, a second GLP1 polypeptide (GLP1B).
Embodiment 87
[0202] The contiguous polypeptide of embodiment 65 comprising:
GLP1A-L1-Fc; or formula (I):
Fc-L1GLP1A. formula (II):
Embodiment 88
[0203] The contiguous polypeptide of embodiment 65 comprising:
GLP1A-L1-Fc-L2-GLP1B. formula (III):
Embodiment 89
[0204] The contiguous polypeptide of any one of embodiments 86 to 88, wherein GLP1B, if present, comprises the same amino acid sequence as GLP1A.
Embodiment 90
[0205] A contiguous polypeptide comprising:
[0206] a) a glucagon-like peptide-1 (GLP1) polypeptide;
[0207] b) a first linker (L1);
[0208] c) an Fc polypeptide (Fc);
[0209] d) a second linker (L2); and
[0210] e) a glucagon polypeptide (Gluc).
Embodiment 91
[0211] The contiguous polypeptide of embodiment 90 comprising:
GLP1-L1-Fc-L2-Gluc; or Formula (IV):
Gluc-L1-Fc-L2-GLP1. Formula (V):
Embodiment 92
[0212] The contiguous polypeptide of any one of embodiments 86 to 91, wherein GLP1A, GLP1, and/or GLP1B, if present, comprises a wild-type GLP1 polypeptide.
Embodiment 93
[0213] The contiguous polypeptide of any one of embodiments 86 to 92, wherein GLP1A, GLP1, and/or GLP1B, if present, comprises a variant GLP1 polypeptide.
Embodiment 94
[0214] The contiguous polypeptide of any one of embodiments 86 to 93, wherein GLP1A, GLP1, and/or GLP1B, if present, comprises an amino acid sequence of SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 98, or SEQ ID NO: 99.
Embodiment 95
[0215] The contiguous polypeptide of any one of embodiments 86 to 94, wherein the glucagon polypeptide comprises a wild-type glucagon polypeptide, optionally comprising the amino acid sequence of SEQ ID NO: 21.
Embodiment 96
[0216] The contiguous polypeptide of any one of embodiments 86 to 95, wherein the glucagon polypeptide is a variant glucagon polypeptide.
Embodiment 97
[0217] The contiguous polypeptide of any one of embodiments 86 to 96, wherein the Fc polypeptide is a human IgG Fc.
Embodiment 98
[0218] The contiguous polypeptide of any one of embodiments 86 to 97, wherein the Fc polypeptide is a human IgG1 Fc, IgG2 Fc, IgG3 Fc, or IgG4 Fc.
Embodiment 99
[0219] The contiguous polypeptide of any one of embodiments 86 to 98, wherein the Fc polypeptide is an Fc of a companion animal species.
Embodiment 100
[0220] The contiguous polypeptide of any one of embodiments 86 to 97 or 99, wherein the Fc polypeptide comprises:
[0221] a) a canine IgG-A Fc, IgG-B Fc, IgG-C Fc, or IgG-D Fc;
[0222] b) an equine IgG1 Fc, IgG2 Fc, IgG3 Fc, IgG4 Fc, IgG5 Fc, IgG6 Fc, or IgG7 Fc; or
[0223] c) a feline IgG1a Fc, IgG1b Fc, or IgG2 Fc.
Embodiment 101
[0224] The contiguous polypeptide of any one of embodiments 86 to 100, wherein the Fc polypeptide is a wild-type IgG Fc polypeptide.
Embodiment 102
[0225] The contiguous polypeptide of any one of embodiments 86 to 100, wherein the Fc polypeptide is a variant IgG Fc polypeptide.
Embodiment 103
[0226] The contiguous polypeptide of any one of embodiments 86 to 102, wherein the Fc polypeptide comprises the polypeptide, the contiguous polypeptide, or the heterodimeric protein of any one of embodiments 1 to 84.
Embodiment 104
[0227] The contiguous polypeptide of any one of embodiments 85 to 102, wherein the contiguous polypeptide has a longer serum half-life than a wild-type GLP1 polypeptide.
Embodiment 105
[0228] The contiguous polypeptide of any one of embodiments 86 to 104, wherein L1 and L2, if present, each independently is a flexible linker.
Embodiment 106
[0229] The contiguous polypeptide of any one of embodiments 86 to 105, wherein the amino acid sequence of L1 and L2, if present, each independently comprises 100%, at least 95%, at least 90%, at least 85% serine and/or glycine amino acid residues.
Embodiment 107
[0230] The contiguous polypeptide of any one of embodiments 86 to 106, wherein the contiguous polypeptide comprises an extension at its C-terminus.
Embodiment 108
[0231] The contiguous polypeptide of any one of embodiments 86 to 107, wherein the contiguous polypeptide comprises a glycine residue, two glycine residues, three glycine residues, four glycine residues, five glycine residues, six glycine residues, seven glycine residues, eight glycine residues, or greater than eight glycine residues at its C-terminus.
Embodiment 109
[0232] The contiguous polypeptide of any one of embodiments 86 to 108, wherein the contiguous polypeptide comprises an amino acid sequence of SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, or SEQ ID NO: 95 at its C-terminus.
Embodiment 110
[0233] The contiguous polypeptide of any one of embodiments 86 to 109, wherein the contiguous polypeptide comprises:
[0234] a) the amino acid sequence of SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; SEQ ID NO: 26; SEQ ID NO: 27; SEQ ID NO: 28; SEQ ID NO: 29; SEQ ID NO: 30; SEQ ID NO: 31; SEQ ID NO: 32; SEQ ID NO: 33; SEQ ID NO: 34; SEQ ID NO: 35; SEQ ID NO: 36; SEQ ID NO: 37; SEQ ID NO: 38; SEQ ID NO: 39; SEQ ID NO: 40; SEQ ID NO: 41; SEQ ID NO: 42; SEQ ID NO: 43; SEQ ID NO: 44; SEQ ID NO: 45; SEQ ID NO: 46; SEQ ID NO: 47, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 103, SEQ ID NO: 104, SEQ ID NO: 105, or SEQ ID NO: 106; or
[0235] b) the amino acid sequence of SEQ ID NO: 52; SEQ ID NO: 53; SEQ ID NO: 54; SEQ ID NO: 55; SEQ ID NO: 56; SEQ ID NO: 57; SEQ ID NO: 58; or SEQ ID NO: 59.
Embodiment 111
[0236] A polypeptide comprising an amino acid sequence of SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; SEQ ID NO: 26; SEQ ID NO: 27; SEQ ID NO: 28; SEQ ID NO: 29; SEQ ID NO: 30; SEQ ID NO: 31; SEQ ID NO: 32; SEQ ID NO: 33; SEQ ID NO: 34; SEQ ID NO: 35; SEQ ID NO: 36; SEQ ID NO: 37; SEQ ID NO: 38; SEQ ID NO: 39; SEQ ID NO: 40; SEQ ID NO: 41; SEQ ID NO: 42; SEQ ID NO: 43; SEQ ID NO: 44; SEQ ID NO: 45; SEQ ID NO: 46; SEQ ID NO: 47, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 52; SEQ ID NO: 53; SEQ ID NO: 54; SEQ ID NO: 55; SEQ ID NO: 56; SEQ ID NO: 57; SEQ ID NO: 58; SEQ ID NO: 59; SEQ ID NO: 103; SEQ ID NO: 104, SEQ ID NO: 105, or SEQ ID NO: 106.
Embodiment 112
[0237] The polypeptide, the heterodimeric protein, or the contiguous polypeptide of any one of the preceding embodiments, wherein the at least one amino acid modification or substitution comprises an amino acid substitution with an amino acid derivative.
Embodiment 113
[0238] An isolated nucleic acid encoding the polypeptide, the heterodimeric protein, or the contiguous polypeptide of any one of the preceding embodiments.
Embodiment 114
[0239] A host cell comprising the nucleic acid of embodiment 113.
Embodiment 115
[0240] A method of producing a polypeptide comprising culturing the host cell of embodiment 114 and isolating the polypeptide.
Embodiment 116
[0241] A pharmaceutical composition comprising the polypeptide, the heterodimeric protein, or the contiguous polypeptide of any one of embodiments 1 to 112, and a pharmaceutically acceptable carrier.
Embodiment 117
[0242] A method of increasing production of cAMP in a cell, the method comprising exposing the cell to the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of embodiments 1 to 112 or 116 under conditions permissive for binding of the polypeptide, heterodimeric protein, or contiguous polypeptide to GLP1R.
Embodiment 118
[0243] The method of embodiment 117, wherein the cell is exposed to the polypeptide, heterodimeric protein, contiguous polypeptide, or the pharmaceutical composition ex vivo.
Embodiment 119
[0244] The method of embodiment 117, wherein the cell is exposed to the polypeptide, heterodimeric protein, contiguous polypeptide, or the pharmaceutical composition in vivo.
Embodiment 120
[0245] The method of any one of embodiments 118 to 119, wherein the cell is a human cell, a canine cell, a feline cell, or an equine cell.
Embodiment 121
[0246] A method of delivering a polypeptide to a subject comprising administering the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of embodiments 1 to 112 or 116 parenterally.
Embodiment 122
[0247] A method of delivering a polypeptide to a subject comprising administering the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of embodiments 1 to 112 or 116 by an intramuscular route, an intraperitoneal route, an intracerobrospinal route, a subcutaneous route, an intra-arterial route, an intrasynovial route, an intrathecal route, or an inhalation route.
Embodiment 123
[0248] A method of treating a subject having diabetes or obesity, the method comprising administering to the subject a therapeutically effective amount of the polypeptide, the heterodimeric protein, the contiguous polypeptide, or the pharmaceutical composition of any one of embodiments 1 to 112 or 116.
Embodiment 124
[0249] The method of embodiment 123, comprising administering insulin, a DPP4 inhibitor, a SGLT2 inhibitor, a biguanides sulfonylureas meglitinide derivative, an alpha-glucosidase inhibitor, a thiazolidinedion (TZD), an amylinomimetic, a bile acid sequestrant, a dopamine agonist.
Embodiment 125
[0250] The method of any one of embodiments 121 to 124, wherein the subject is a human subject.
Embodiment 126
[0251] The method of any one of embodiments 121 to 124, wherein the subject is a companion animal species.
Embodiment 127
[0252] The method of embodiment 126, wherein the companion animal species is canine, equine, or feline.
BRIEF DESCRIPTION OF THE DRAWINGS
[0253] FIG. 1 shows an alignment of canine IgG-A, B, C, and D Fc sequences. The boxes indicate the regions likely in contact with Protein A.
[0254] FIG. 2A shows an SDS-PAGE analysis of GLP1-G8/GLP-2G_III_WTfeIgG2 (SEQ ID NO: 23; "GLP1 A variant" in this figure) and GLP1-G8_I_WTfeIgG2 (SEQ ID NO: 24; "GLP1 B variant" in this figure) having wild-type feline IgG2 hinge with one disulfide bond in the absence and presence of reducing agent (DTT).
[0255] FIG. 2B shows an SDS-PAGE analysis of GLP1-G8/GLP-2G_III_VARfeIgG2 (SEQ ID NO: 25; "GLP1 MA variant" in this figure) of GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26; "GLP1 MB variant" in this figure) having variant feline IgG2 hinge with two disulfide bonds in the absence and presence of reducing agent (DTT).
[0256] FIG. 3 shows a cAMP CHO-K1 GLP1R Bioassay to evaluate activity of GLP1-G8/GLP1-2G_III_VARfeIgG2 (SEQ ID NO: 25) and GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) compared to controls (GLP1 (7-37) and Extendin-4).
[0257] FIG. 4 shows a cell-based bioassay to evaluate activity of GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) ("GLP1-B" in this figure) after 1 year of storage with CHOK1-GLP1R cells and cAMP-glo (n=2).
[0258] FIG. 5 shows a Western Blot analysis of GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) ("GLP1B" in this figure) after incubation in serum for 24 hours at 37.degree. C. (lane 1), in PBS for 24 hours at 37.degree. C. (lane 2), in PBS for 24 hours at 4.degree. C. (lane 9). A mouse anti-GLP1 antibody was used.
[0259] FIG. 6 is a plot of GLP1-G8_I_VARfeIgG2 (SEQ ID NO:26) concentration in the serum over time after subcutaneous administration to 5 cats, as measured by quantitative ELISA.
[0260] FIG. 7 shows a plot of GLP1-G8_I_VARfeIgG2 (SEQ ID NO:26) concentration in the serum over time after subcutaneous administration to 5 cats, as measured by cell-based activity assay. The mean AUC from 0 to 168 hours was about 840 .mu.g(h)/mL and the mean t.sub.1/2 was 36 hours.
DESCRIPTION OF THE SEQUENCES
TABLE-US-00001
[0261] TABLE 1 Table 1 provides a listing of exemplary sequences referenced herein. Description of the Sequences SEQ ID NO: SEQUENCE DESCRIPTION 1 PVPEPLGGPSVLIFPPKPKDILRITRIPEVIC Exemplary wild-type canine VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIEHQDWLTGKEFKCRVNH Protein A- IDLPSPIERTISKARGRAHKPSVYVLPPSPKE C1q- LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE CD16- PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 2 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary wild-type canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16+ ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 107 PKRENGRVPRPPDCPKCPAPEMLGGPSVFIFPP Exemplary wild-type canine KPKDILLIARTPEVICVVVDLDPEDPEVQISW IgG-B Fc with hinge FVDGKQMQTAKTQPREEQFNGTYRVVSVLPIG Protein A+ HQDWLKGKQFTCKVNNKALPSPIERTISKARG C1q+ QAHQPSVYVLPPSREELSKNIVSLICLIKDFF CD16+ PPDIDVEWQSNGQQEPESKYRTTPPQLDEDGS YFLYSKLSVDKSRWQRGDTFICAVMHEALHNH YTQESLSHSPGK 3 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary wild-type canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A- KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16+ ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 108 AKECECKCNCNNCPCPGCGLLGGPSVFIFPPK Exemplary wild-type canine PKDILVTARTPTVTCVVVDLDPENPEVQISWF IgG-C Fc with hinge VDSKQVQTANTQPREEQSNGTYRVVSVLPIGH Protein A- QDWLSGKQFKCKVNNKALPSPIEEIISKTPGQ C1q+ AHQPNVYVLPPSRDEMSKNIVTLICLVKDFFP CD16+ PEIDVEWQSNGQQEPESKYRMTPPQLDEDGSY FLYSKLSVDKSRWQRGDTFICAVMHEALHNHY TQISLSHSPGK 4 PVPESLGGPSVFIFPPKPKDILRITRIPEITC Exemplary wild-type canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIEHQDWLIGKEFKCRVNH Protein A- IGLPSPIERTISKARGQAHQPSVYVLPPSPKE C1q- LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE CD16- PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 5 PVPEPLGGPSVLIFPPKPKDTLLIARTPEVIC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHIAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIGHQDWLIGKEFKCRVNH C1q- IDLPSPIERTISKARGRAHKPSVYVLPPSPKE Protein A+ LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE I(21)T PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ R(23)L QGDPFTCAVMHEALHNHYTDLSLSHSPGK T(25)A E(80)G T(205)A Q(207)H 6 PGCGLLGGPSVFIFPPKPKDTLLIARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN C1q+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE Protein A+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP I(21)T ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR V(23)L GDTFICAVMHEALHNHYTQISLSHSPGK T(24)I 7 PVPESLGGPSVFIFPPKPKDTLLIARTPEITC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIGHQDWLTGKEFKCRVNH C1q- IGLPSPIERTISKARGQAHQPSVYVLPPSPKE Protein A+ LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE I(21)T PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ R(23)L QGDTFTCAVMHEALHNHYTDLSLSHSPGK T(25)A E(80)G Q(207)H 8 PVPEPLGGPSVLIFPPKPKDILRITRIPEVIC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIEHQDWLTGKEFKCRVNH Heterodimer chain 1 T(138)Y IDLPSPIERTISKARGRAHKPSVYVLPPSPKE LSSSDTVSIYCLIKDFYPPDIDVEWQSNGQQE PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 9 PVPEPLGGPSVLIFPPKPKDILRITRIPEVIC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIEHQDWLTGKEFKCRVNH Heterodimer chain 2 IDLPSPIERTISKARGRAHKPSVYVLPPSPKE Y(181)T LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE PERKHRMTPPQLDEDGSYFLTSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 10 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Heterodimer chain 1 T(137)Y KALPSPIERTISKARGQAHQPSVYVLPPSREE LSKNTVSLYCLIKDFFPPDIDVEWQSNGQQEP ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 11 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Heterodimer chain 2 KALPSPIERTISKARGQAHQPSVYVLPPSREE Y(180)T LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP ESKYRTTPPQLDEDGSYFLTSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 12 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Heterodimer chain 1 T(137)Y KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE MSKNTVTLYCLVKDFFPPEIDVEWQSNGQQEP ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 13 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Heterodimer chain 2 KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE Y(180)T MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP ESKYRMTPPQLDEDGSYFLTSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 14 PVPESLGGPSVFIFPPKPKDILRITRITEITC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIEHQDWLTGKEFKCRVNH Heterodimer chain 1 T(138)Y IGLPSPIERTISKARGQAHQPSVYVLPPSPKE LSSSDTVTLYCLIKDFFPPEIDVEWQSNGQPE PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 15 PVPESLGGPSVFIFPPKPKDILRITRITEITC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIEHQDWLTGKEFKCRVNH Heterodimer chain 2 IGLPSPIERTISKARGQAHQPSVYVLPPSPKE Y(181)T LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE PESKYHTTAPQLDEDGSYFLTSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 109 PVPEPLGGPSVLIFPPKPKDILRITRTPEVIC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIEHQDWLTGKEFKCRVNH Heterodimer chain 3 IDLPSPIERTISKARGRAHKPSVYVLPPSPKE T(138)W LSSSDTVSIWCLIKDFYPPDIDVEWQSNGQQE PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 110 PVPEPLGGPSVLIFPPKPKDILRITRITEVIC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIEHQDWLTGKEFKCRVNH Heterodimer chain 4 IDLPSPIERTISKARGRAHKPSVYVLPPSPKE T(138)S LSSSDTVSISCAIKDFYPPDIDVEWQSNGQQE L(140)A PERKHRMTPPQLDEDGSYFLTSKLSVDKSRWQ Y(181)T QGDPFTCAVMHETLQNHYTDLSLSHSPGK 111 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Heterodimer chain 3 KALPSPIERTISKARGQAHQPSVYVLPPSREE T(137)W LSKNTVSLWCLIKDFFPPDIDVEWQSNGQQEP ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 112 PAPEMLGGPSVFIFPPKPKDTLLIARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Heterodimer chain 4 KALPSPIERTISKARGQAHQPSVYVLPPSREE T(137)S LSKNTVSLSCAIKDFFPPDIDVEWQSNGQQEP L(139)A ESKYRTTPPQLDEDGSYFLTSKLSVDKSRWQR Y(180)T GDTFICAVMHEALHNHYTQESLSHSPGK 113 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Heterodimer chain 3 KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE T(137)W MSKNTVTLWCLVKDFFPPEIDVEWQSNGQQEP ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 114 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Heterodimer chain 4 KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE T(137)S MSKNTVTLSCAVKDFFPPEIDVEWQSNGQQEP L(139)A ESKYRMTPPQLDEDGSYFLTSKLSVDKSRWQR Y(180)T GDTFICAVMHEALHNHYTQISLSHSPGK 115 PVPESLGGPSVFIFPPKPKDILRITRITEITC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIEHQDWLIGKEEKCRVNH Heterodimer chain 3 IGLPSPIERTISKARGQAHQPSVYVLPPSPKE T(138)W LSSSDTVTLWCLIKDFFPPEIDVEWQSNGQPE PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 116 PVPESLGGPSVFIFPPKPKDILRITRITEITC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIEHQDWLIGKEEKCRVNH Heterodimer chain 4 IGLPSPIERTISKARGQAHQPSVYVLPPSPKE T(138)S LSSSDTVTLSCAIKDFFPPEIDVEWQSNGQPE L(140)A PESKYHTTAPQLDEDGSYFLTSKLSVDKSRWQ Y(181)T QGDTFTCAVMHEALQNHYTDLSLSHSPGK 16 PKTASTIESKTGEGPKCPVPEIPGAPSVFIFP Exemplary wild-type feline PKPKDTLSISRTPEVICLVVDLGPDDSNVQIT IgG2 Fc WFVDNTEMHTAKTRPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK C1q- GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 117 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary wild-type feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1a Fc WFVDNTQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK C1q+ GQPHEPQVYVLPPAQEELSENKVSVTCLIKSF HPPDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 118 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary wild-type feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1b Fc WFVDNTQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK C1q+ GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF YPSDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 119 PKTASTIESKTGEGPKCPVPEIPGAPSVFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSNVQIT IgG2 Fc WFVDNTEMHTAKTRPREEQFNSTYRVVSVLPI Heterodimer chain 1 LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK T(154)W GQPHEPQVYVLPPTQEELSENKVSVWCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK
120 PKTASTIESKTGEGPKCPVPEIPGAPSVFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSNVQIT IgG2 Fc WFVDNTEMHTAKTRPREEQFNSTYRVVSVLPI Heterodimer chain 2 LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK T(154)S GQPHEPQVYVLPPTQEELSENKVSVSCAIKGF L(156)A HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG Y(197)T TYFLTSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 121 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1a Fc WFVDNTQVYTAKTSPREEQFNSTYRVVSVLPI Heterodimer chain 1 LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK T(154)W GQPHEPQVYVLPPAQEELSENKVSVWCLIKSF HPPDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 122 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1a Fc WFVDNTQVYTAKTSPREEQFNSTYRVVSVLPI Heterodimer chain 2 LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK T(154)S GQPHEPQVYVLPPAQEELSENKVSVSCAIKSF L(156)A HPPDIAVEWEITGQPEPENNYRTTPPQLDSDG Y(197)T TYFVTSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 123 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1b Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Heterodimer chain 1 LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK T(154)W GQPHEPQVYVLPPAQEELSENKVSVWCLIEGF YPSDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 124 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1b Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Heterodimer chain 2 LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK T(154)S GQPHEPQVYVLPPAQEELSENKVSVSCAIEGF L(156)A YPSDIAVEWEITGQPEPENNYRTTPPQLDSDG Y(197)T TYFLTSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 17 PKTASTIESKTGECPKCPVPEIPGAPSVFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSNVQIT IgG2 Fc WFVDNTEMHTAKTRPREEQENSTYRVVSVLPI Hinge Cys LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK G(14)C GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 125 RKTDHPPGPKPCDCPKCPPPEMLGGPSVFIFP Exemplary variant feline PKPKDTLSISRTPEVTCLVVDLGPDDSNVQIT IgG2 Fc with feline IgG1 WFVDNTEMHTAKTRPREEQENSTYRVVSVLPI hinge LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 126 PKTASTIESKTGEGPPCPVPEIPGAPSVFIFP Exemplary variant feline PKPKDTLSISRTPEVTCLVVDLGPDDSNVQIT IgG2 Fc with modified hinge WFVDNTEMHTAKTRPREEQENSTYRVVSVLPI K(16)P LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 127 RKTDHPPGPKPCDCPPCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT IgG1a Fc with modified hinge WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI K(16)P LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK GQPHEPQVYVLPPAQEELSENKVSVTCLIKSF HPPDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 128 RKTDHPPGPKPCDCPPCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT IgG1b Fc with modified WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI hinge LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK K(16)P GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF YPSDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 18 DMSKCPKCPAPELLGGPSVFIFPPNPKDALMI Exemplary variant equine Fc SRTPVVTCVVVNLSDQYPDVQFSWYVDNTEVH IgG2 (with equine IgG1 hinge) SAITKQREAQFNSTYRVVSVLPIQHQDWLSGK Protein A+ EFKCSVTNVGVPQPISRAISRGKGPSRVPQVY C1q- VLPPHPDELAKSKVSVTCLVKDFYPPDISVEW QSNRWPELEGKYSTTPAQLDGDGSYFLYSKLS LETSRWQQVESFICAVMHEALHNHFIKTDISE SLGK 19 DMSKCPKCPAPELLGGPSVFIFPPNPKDTLMI Exemplary variant equine SRTPVVTCVVVNLSDQYPDVQFSWYVDNTEVH IgG2 Fc (with equine IgG1 hinge) SAITKQREAQFNSTYRVVSVLPIQHQDWLSGK C1q- EFKCSVTNVGVPQPISRAISRGKGPSRVPQVY Protein A+ VLPPHPDELAKSKVSVTCLVKDFYPPDISVEW A(29)T QSNRWPELEGKYSTTPAQLDGDGSYFLYSKLS F(217)Y LETSRWQQVESFICAVMHEALHNHYTKTDISE SLGK 20 HXEGTFTSDVSSYLEGQAAKEFIAWLVKG Exemplary variant GLP1 (7-35) X8 may be G or S 21 HSQGTFTSDYSKYLDSRRAQDFVQWLMNT Glucagon (Gluc) 22 HGEGTFTSDLSKQMEEEAVRLFIEWLKNGGPS Extendin-4 SGAPPPS 23 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8/GLP-2G_III_ SGGGGSGGGGSGGGGSPKTASTIESKTGEGPK WTfeIgG2 CPVPEIPGAPSVFIFPPKPKDTLSISRTPEVT CLVVDLGPDDSNVQITWFVDNTEMHTAKTRPR EEQFNSTYRVVSVLPILHQDWLKGKEFKCKVN SKSLPSAMERTISKAKGQPHEPQVYVLPPTQE ELSENKVSVTCLIKGFHPPDIAVEWEITGQPE PENNYQTTPPQLDSDGTYFLYSRLSVDRSHWQ RGNTYTCSVSHEALHSHHTQKSLTQSPGKGGG GSGGGGHAEGTFTSDVSSYLEGQAAKEFIAWL VKGGG 24 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8_I_WTfeIgG2 SGGGGSGGGGSGGGGSPKTASTIESKTGEGPK CPVPEIPGAPSVFIFPPKPKDTLSISRTPEVT CLVVDLGPDDSNVQITWFVDNTEMHTAKTRPR EEQFNSTYRVVSVLPILHQDWLKGKEFKCKVN SKSLPSAMERTISKAKGQPHEPQVYVLPPTQE ELSENKVSVTCLIKGFHPPDIAVEWEITGQPE PENNYQTTPPQLDSDGTYFLYSRLSVDRSHWQ RGNTYTCSVSHEALHSHHTQKSLTQSPGK 25 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8/GLP1-2G_III_ SGGGGSGGGGSGGGGSPKTASTIESKTGECPK VARfeIgG2 CPVPEIPGAPSVFIFPPKPKDTLSISRTPEVT CLVVDLGPDDSNVQITWFVDNTEMHTAKTRPR EEQFNSTYRVVSVLPILHQDWLKGKEFKCKVN SKSLPSAMERTISKAKGQPHEPQVYVLPPTQE ELSENKVSVTCLIKGFHPPDIAVEWEITGQPE PENNYQTTPPQLDSDGTYFLYSRLSVDRSHWQ RGNTYICSVSHEALHSHHTQKSLTQSPGKGGG GSGGGGHAEGTFTSDVSSYLEGQAAKEFIAWL VKGGG 26 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8_I_VARfeIgG2 SGGGGSGGGGSGGGGSPKTASTIESKTGECPK CPVPEIPGAPSVFIFPPKPKDTLSISRTPEVT CLVVDLGPDDSNVQITWFVDNTEMHTAKTRPR EEQFNSTYRVVSVLPILHQDWLKGKEFKCKVN SKSLPSAMERTISKAKGQPHEPQVYVLPPTQE ELSENKVSVTCLIKGFHPPDIAVEWEITGQPE PENNYQTTPPQLDSDGTYFLYSRLSVDRSHWQ RGNTYTCSVSHEALHSHHTQKSLTQSPGK 27 HSEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-S8/GLP1-3G_III_ GGGGSGGGSGGGSGGGSPKTASTIESKTGEGP WTfeIgG2 KCPVPEIPGAPSVFIFPPKPKDTLSISRTPEV TCLVVDLGPDDSNVQITWFVDNTEMHTAKTRP REEQFNSTYRVVSVLPILHQDWLKGKEFKCKV NSKSLPSAMERTISKAKGQPHEPQVYVLPPTQ EELSENKVSVTCLIKGFHPPDIAVEWEITGQP EPENNYQTTPPQLDSDGTYFLYSRLSVDRSHW QRGNTYTCSVSHEALHSHHTQKSLTQSPGKGG GGSGGGGHAEGTFTSDVSSYLEGQAAKEFTAW LVKGGGG 28 HSEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-S8_I_WTfeIgG2 GGGGSGGGSGGGSGGGSPKTASTIESKTGEGP KCPVPEIPGAPSVFIFPPKPKDTLSISRTPEV TCLVVDLGPDDSNVQITWFVDNTEMHTAKTRP REEQFNSTYRVVSVLPILHQDWLKGKEFKCKV NSKSLPSAMERTISKAKGQPHEPQVYVLPPTQ EELSENKVSVTCLIKGFHPPDIAVEWEITGQP EPENNYQTTPPQLDSDGTYFLYSRLSVDRSHW QRGNTYTCSVSHEALHSHHTQKSLTQSPGK 29 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8/GLP1-3G_III_ GGGGSGGGSGGGSGGGSPKESTSKCISPCPVP VARcaIgGD ESLGGPSVFIFPPKPKDTLLIARTPEITCVVL DLGREDPEVQISWFVDGKEVHTAKTQPREQQF NSTYRVVSVLPIGHQDWLIGKEEKCRVNHIGL PSPIERTISKARGQAHQPSVYVLPPSPKELSS SDTVTLTCLIKDFFPPEIDVEWQSNGQPEPES KYHTTAPQLDEDGSYFLYSKLSVDKSRWQQGD TFTCAVMHEALHNHYTDLSLSHSPGKGGGGSG GGGHAEGTFTSDVSSYLEGQAAKEFIAWLVKG GGG 30 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8_I_VARcaIgGD GGGGSGGGSGGGSGGGSPKESTCKCISPCPVP ESLGGPSVFIFPPKPKDTLLIARTPEITCVVL DLGREDPEVQISWFVDGKEVHTAKTQPREQQF NSTYRVVSVLPIGHQDWLIGKEEKCRVNHIGL PSPIERTISKARGQAHQPSVYVLPPSPKELSS SDTVTLTCLIKDFFPPEIDVEWQSNGQPEPES KYHTTAPQLDEDGSYFLYSKLSVDKSRWQQGD TFTCAVMHEALHNHYTDLSLSHSPGK 31 HSEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-88/GLP1-3G_III_ GGGGSGGGSGGGSGGGSPKESTSKCISPCPVP VARcaIgGD ESLGGPSVFIFPPKPKDTLLIARTPEITCVVL DLGREDPEVQISWFVDGKEVHTAKTQPREQQF NSTYRVVSVLPIGHQDWLTGKEFKCRVNHIGL PSPIERTISKARGQAHQPSVYVLPPSPKELSS SDTVTLTCLIKDFFPPEIDVEWQSNGQPEPES KYHTTAPQLDEDGSYFLYSKLSVDKSRWQQGD TFTCAVMHEALHNHYTDLSLSHSPGKGGGGSG GGGHAEGTFTSDVSSYLEGQAAKEFIAWLVKG GGG 32 HSEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-S8_I_VARcaIgGD GGGGSGGGSGGGSGGGSPKESTCKCISPCPVP ESLGGPSVFIFPPKPKDTLLIARTPEITCVVL DLGREDPEVQISWFVDGKEVHTAKTQPREQQF NSTYRVVSVLPIGHQDWLTGKEFKCRVNHIGL PSPIERTISKARGQAHQPSVYVLPPSPKELSS SDTVTLTCLIKDFFPPEIDVEWQSNGQPEPES KYHTTAPQLDEDGSYFLYSKLSVDKSRWQQGD TFTCAVMHEALHNHYTDLSLSHSPGK 33 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8/GLPL1-3G_III_ GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG VAReqIgG2 GPSVFIFPPNPKDTLMISRTPVVICVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VTCLVKDFYPPDISVEWQSNRWPELEGKYSTT PAQLDGDGSYFLYSKLSLETSRWQQVESFTCA VMHEALHNHYTKTDISESLGKGGGGSGGGGHA EGTFTSDVSSYLEGQAAKEFIAWLVKGGGG 34 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8_I_VAReqIgG2 GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG GPSVFIFPPNPKDTLMISRTPVVICVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VTCLVKDFYPPDISVEWQSNRWPELEGKYSTT PAQLDGDGSYFLYSKLSLETSRWQQVESFTCA VMHEALHNHYTKTDISESLGK 35 HSEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-88/GLP1-3G_III_ GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG VAReqIgG2 GPSVFIFPPNPKDTLMISRTPVVICVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY
RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VICLVKDFYPPDISVEWQSNRWPELEGKYSIT PAQLDGDGSYFLYSKLSLETSRWQQVESFTCA VMHEALHNHYTKTDISESLGKGGGGSGGGGHA EGTFTSDVSSYLEGQAAKEFIAWLVKGGGG 36 HSEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-S8_I_VAReqIgG2 GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG GPSVFIFPPNPKDTLMISRTPVVTCVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VTCLVKDFYPPDISVEWQSNRWPELEGKYSTT PAQLDGDGSYFLYSKLSLETSRWQQVESFICA VMHEALHNHYTKTDISESLGK 37 MAVLGLLFCLVTFPSCVLSHSEGTFTSDVSSY ssGLP1-S8/GLP1-3G_III_ LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS WTfeIgG2 GGGSPKTASTIESKTGEGPKCPVPEIPGAPSV FIFPPKPKDTLSISRTPEVTCLVVDLGPDDSN VQITWFVDNTEMHTAKTRPREEQFNSTYRVVS VLPILHQDWLKGKEFKCKVNSKSLPSAMERTI SKAKGQPHEPQVYVLPPTQEELSENKVSVTCL IKGFHPPDIAVEWEITGQPEPENNYQTTPPQL DSDGTYFLYSRLSVDRSHWQRGNTYTCSVSHE ALHSHHTQKSLTQSPGKGGGGSGGGGHAEGTF TSDVSSYLEGQAAKEFIAWLVKGGGG 38 MAVLGLLFCLVTFPSCVLSHGEGTFTSDVSSY ssGLP1-G8/GLP1-2G_III_ LEGQAAKEFIAWLVKGGGGSGGGGSGGGGSGG VARfeIgG2 GGSPKTASTIESKTGECPKCPVPEIPGAPSVF IFPPKPKDTLSISRTPEVICLVVDLGPDDSNV QITWFVDNTEMHTAKTRPREEQFNSTYRVVSV LPILHQDWLKGKEFKCKVNSKSLPSAMERTIS KAKGQPHEPQVYVLPPTQEELSENKVSVTCLI KGFHPPDIAVEWEITGQPEPENNYQTTPPQLD SDGTYFLYSRLSVDRSHWQRGNTYTCSVSHEA LHSHHTQKSLTQSPGKGGGGSGGGGHAEGTFT SDVSSYLEGQAAKEFIAWLVKGGG 39 MAVLGLLFCLVTFPSCVLSHGEGTFTSDVSSY ssGLP1-G8_I_VARfeIgG2 LEGQAAKEFIAWLVKGGGGSGGGGSGGGGSGG GGSPKTASTIESKTGECPKCPVPEIPGAPSVF IFPPKPKDTLSISRTPEVICLVVDLGPDDSNV QITWFVDNTEMHTAKTRPREEQFNSTYRVVSV LPILHQDWLKGKEFKCKVNSKSLPSAMERTIS KAKGQPHEPQVYVLPPTQEELSENKVSVTCLI KGFHPPDIAVEWEITGQPEPENNYQTTPPQLD SDGTYFLYSRLSVDRSHWQRGNTYTCSVSHEA LHSHHTQKSLTQSPGK 40 MAVLGLLFCLVTFPSCVLSHSEGTFTSDVSSY ssGLP1-S8_I_WTfeIgG2 LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS GGGSPKTASTIESKTGEGPKCPVPEIPGAPSV FIFPPKPKDTLSISRTPEVTCLVVDLGPDDSN VQITWFVDNTEMHTAKTRPREEQFNSTYRVVS VLPILHQDWLKGKEFKCKVNSKSLPSAMERTI SKAKGQPHEPQVYVLPPTQEELSENKVSVTCL IKGFHPPDIAVEWEITGQPEPENNYQTTPPQL DSDGTYFLYSRLSVDRSHWQRGNTYTCSVSHE ALHSHHTQKSLTQSPGK 41 MAVLGLLFCLVTFPSCVLSHGEGIFTSDVSSY ssGLP1-G8/GLP1-2G_III_ LEGQAAKEFIAWLVKGGGGSGGGGSGGGGSGG VARcaIgGD GGSPKESTCKCISPCPVPESLGGPSVFIFPPK PKDTLLIARTPEITCVVLDLGREDPEVQISWF VDGKEVHTAKTQPREQQFNSTYRVVSVLPIGH QDWLTGKEFKCRVNHIGLPSPIERTISKARGQ AHQPSVYVLPPSPKELSSSDTVTLTCLIKDFF PPEIDVEWQSNGQPEPESKYHTTAPQLDEDGS YFLYSKLSVDKSRWQQGDTFTCAVMHEALHNH YTDLSLSHSPGKGGGGSGGGGHAEGTFTSDVS SYLEGQAAKEFIAWLVKGGG 42 MAVLGLLFCLVTFPSCVLSHGEGIFTSDVSSY ssGLP1-G8/GLP1-2G_III_ LEGQAAKEFIAWLVKGGGGSGGGGSGGGGSGG VARcaIgGD GGSPKESTSKCISPCPVPESLGGPSVFIFPPK PKDTLLIARTPEITCVVLDLGREDPEVQISWF VDGKEVHTAKTQPREQQFNSTYRVVSVLPIGH QDWLTGKEFKCRVNHIGLPSPIERTISKARGQ AHQPSVYVLPPSPKELSSSDTVTLTCLIKDFF PPEIDVEWQSNGQPEPESKYHTTAPQLDEDGS YFLYSKLSVDKSRWQQGDTFTCAVMHEALHNH YTDLSLSHSPGKGGGGSGGGGHAEGTFTSDVS SYLEGQAAKEFIAWLVKGGG 43 MAVLGLLFCLVTFPSCVLSHSEGTFTSDVSSY ssGLP1-S8/GLP1-3G_III_ LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS VARcaIgGD GGGSPKESTSKCISPCPVPESLGGPSVFIFPP KPKDTLLIARTPEITCVVLDLGREDPEVQISW FVDGKEVHTAKTQPREQQFNSTYRVVSVLPIG HQDWLTGKEFKCRVNHIGLPSPIERTISKARG QAHQPSVYVLPPSPKELSSSDTVTLTCLIKDF FPPEIDVEWQSNGQPEPESKYHTTAPQLDEDG SYFLYSKLSVDKSRWQQGDTFTCAVMHEALHN HYTDLSLSHSPGKGGGGSGGGGHAEGTFTSDV SSYLEGQAAKEFIAWLVKGGGG 44 MAVLGLLFCLVTFPSCVLSHGEGIFTSDVSSY ssGLP1-G8/GLP1-3G_III_ LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS VAReqIgG2 GGGSDMSKCPKCPAPELLGGPSVFIFPPNPKD TLMISRIPVVICVVVNLSDQYPDVQFSWYVDN TEVHSAITKQREAQFNSTYRVVSVLPIQHQDW LSGKEFKCSVTNVGVPQPISRAISRGKGPSRV PQVYVLPPHPDELAKSKVSVTCLVKDFYPPDI SVEWQSNRWPELEGKYSTTPAQLDGDGSYFLY SKLSLETSRWQQVESFTCAVMHEALHNHYTKT DISESLGKGGGGSGGGGHAEGTFTSDVSSYLE GQAAKEFIAWLVKGGGG 45 MAVLGLLFCLVTFPSCVLSHGEGIFTSDVSSY ssGLP1-G8_I_VAReqIgG2 LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS GGGSSVPKPQCPPYTHSKFLGGPSVFIFPPNP KDTLMISRTPVVICVVVNLSDQYPDVQFSWYV DNTEVHSAITKQREAQFNSTYRVVSVLPIQHQ DWLSGKEFKCSVTNVGVPQPISRAISRGKGPS RVPQVYVLPPHPDELAKSKVSVTCLVKDFYPP DISVEWQSNRWPELEGKYSITPAQLDGDGSYF LYSKLSLETSRWQQVESFTCAVMHEALHNHYT KTDISESLGK 46 MAVLGLLFCLVTFPSCVLSHSEGTFTSDVSSY ssGLP1-S8/GLP1-3G_III_ LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS VAReqIgG2 GGGSDMSKCPKCPAPELLGGPSVFIFPPNPKD TLMISRIPVVICVVVNLSDQYPDVQFSWYVDN TEVHSAITKQREAQFNSTYRVVSVLPIQHQDW LSGKEFKCSVTNVGVPQPISRAISRGKGPSRV PQVYVLPPHPDELAKSKVSVICLVKDFYPPDI SVEWQSNRWPELEGKYSTTPAQLDGDGSYFLY SKLSLETSRWQQVESFTCAVMHEALHNHYTKT DISESLGKGGGGSGGGGHAEGTFTSDVSSYLE GQAAKEFIAWLVKGGGG 47 MAVLGLLFCLVTFPSCVLSHSEGIFTSDVSSY ssGLP1-S8_I_VAReqIgG2 LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS GGGSSVPKPQCPPYTHSKFLGGPSVFIFPPNP KDTLMISRTPVVICVVVNLSDQYPDVQFSWYV DNTEVHSAITKQREAQFNSTYRVVSVLPIQHQ DWLSGKEFKCSVTNVGVPQPISRAISRGKGPS RVPQVYVLPPHPDELAKSKVSVICLVKDFYPP DISVEWQSNRWPELEGKYSITPAQLDGDGSYF LYSKLSLETSRWQQVESFTCAVMHEALHNHYT KTDISESLGK 48 MGLVAPVVLLHQDDEEHGQDEGPEDGSGYLLG Feline glucagon-like peptide TLTRFSSDFDSAPEVILAPDDQLQLPHPSSRE 1 receptor (GLP1R) NFWARTGLCAESFLLRPVGPVGPVMGWSEGFH KRNSRQEFLRRRLFAGGLCAASTQESRNRCSS RGCKSSPADCPELDRTQHLGNSVGPIQAAHQE LALGAGGPGDECQCCSVSNSLFIPEPQSTCPY NGYTSWPLEGNLRVACAPPPPPPARTLFGGSR RGAVDKKAGGGNRSPGGGAGTGEFGAPGAGGG LGRRPEVGAWTAAEGTNPADLASSPPPPSTRP PAAPRPPCADFCAASPQTTFPIPSPRRPLPAS GGATVSLSETVQKWREYRHQCQRFLTEAPPPA TGLFCNRTFDEYACWPDGLPGSFVNVSCPWYL PWASSVLQGHVYRFCTAEGLWLRQDNSSLPWR NLSECEESKRGERSSPEEQLLSFSIIYTVGYT LSFSALVIASAILLSFRHLHCTRNYIHLNLFA SFILRALSVFIRDAVLKWMYSTAPQQHQWDGL LSYQDSLGCRLVFLLMQYCVAANYYWLLVEGV YLYILLAFSVESEQRIFRLYLSIGWGVPLLFV IWGIVKYLYEDEGCWTRNSNMNYWLIIRLPIL FAIGVNFLIFVRVICIVVSKLKANLMCKTDIK CRLAKSTLTLIPLLGTHEVVFAFVMDEHARGT LRFIKLFTELSFTSFQGLMVAILYCFVNNEVQ MEFRRSWERWRLKHLHIQRDSSMKPLKCPTSS LTSGGTVGSSVYAASCQASCS 49 RPLPASGGATVSLSETVQKWREYRHQCQRFLT Mature feline glucagon-like EAPPPATGLFCNRTFDEYACWPDGLPGSFVNV peptide 1 receptor-N-terminal SCPWYLPWASSVLQGHVYRFCTAEGLWLRQDN domain (GLP1R-N) SSLPWRNLSECEESKRGERSSPEEQLLSFS 50 METDTLLLWVLLLWVPGSTGRPLPASGGATVS ssFeGLP1R-N_huFc_ LSETVQKWREYRHQCQRFLTEAPPPATGLFCN PolyHis RTFDEYACWPDGLPGSFVNVSCPWYLPWASSV LQGHVYRFCTAEGLWLRQDNSSLPWRNLSECE ESKRGERSSPEEQLLSFSGSENLYFQGPKSCD KTHTCPPCPAPELLGGPSVFLFPPKPKDTLMI SRIPEVTCVVVDVSHEDPEVKFNWYVDGVEVH NAKTKPREEQYNSTYRVVSVLTVLHQDWLNGK EYKCKVSNKALPAPIEKTISKAKGQPREPQVY TLPPSRDELTKNQVSLTCLVKGFYPSDIAVEW ESNGQPENNYKTTPPVLDSDGSFFLYSKLTVD KSRWQQGNVFSCSVMHEALHNHYTQKSLSLSP GKHHHHHH 51 METDTLLLWVLLLWVPGSTGRPLPASGGATVS ssFeGLP1R-N_PolyHis LSETVQKWREYRHQCQRFLTEAPPPATGLFCN RTFDEYACWPDGLPGSFVNVSCPWYLPWASSV LQGHVYRFCTAEGLWLRQDNSSLPWRNLSECE ESKRGERSSPEEQLLSFSGGGSHHHHHH 52 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8/Gluc-3G_IV_ SGGGGSGGGGSGGGGSPKTASTIESKTGEGPK WTfeIgG2 CPVPEIPGAPSVFIFPPKPKDTLSISRTPEVT CLVVDLGPDDSNVQITWFVDNTEMHTAKTRPR EEQFNSTYRVVSVLPILHQDWLKGKEFKCKVN SKSLPSAMERTISKAKGQPHEPQVYVLPPTQE ELSENKVSVTCLIKGFHPPDIAVEWEITGQPE PENNYQTTPPQLDSDGTYFLYSRLSVDRSHWQ RGNTYICSVSHEALHSHHTQKSLTQSPGKGGG GSGGGGHSQGTFTSDYSKYLDSRRAQDFVQWL MNTGGG 53 HSQGTFTSDYSKYLDSRRAQDFVQWLMNTGGG Gluc/GLP1-2G_V_ GSGGGGSGGGGSGGGGSPKTASTIESKTGEGP WTfeIgG2 KCPVPEIPGAPSVFIFPPKPKDTLSISRTPEV TCLVVDLGPDDSNVQITWFVDNTEMHTAKTRP REEQFNSTYRVVSVLPILHQDWLKGKEFKCKV NSKSLPSAMERTISKAKGQPHEPQVYVLPPTQ EELSENKVSVTCLIKGFHPPDIAVEWEITGQP EPENNYQTTPPQLDSDGTYFLYSRLSVDRSHW QRGNTYTCSVSHEALHSHHTQKSLTQSPGKGG GGSGGGGHAEGTFTSDVSSYLEGQAAKEFIAW LVKGGG 54 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8/Glu-4G_IV_ GGGGSGGGSGGGSGGGSPKESTSKCISPCPVP VARcaIgGD ESLGGPSVFIFPPKPKDTLLIARTPEITCVVL DLGREDPEVQISWFVDGKEVHTAKTQPREQQF NSTYRVVSVLPIGHQDWLTGKEFKCRVNHIGL PSPIERTISKARGQAHQPSVYVLPPSPKELSS SDTVTLTCLIKDFFPPEIDVEWQSNGQPEPES KYHTTAPQLDEDGSYFLYSKLSVDKSRWQQGD TFTCAVMHEALHNHYTDLSLSHSPGKGGGGSG GGGHSQGTFTSDYSKYLDSRRAQDEVQWLMNT GGGG 55 HSQGTFTSDYSKYLDSRRAQDFVQWLMNTGAG Gluc/GLP1-3G_V_ GGGGSGGGSGGGSGGGSPKESTSKCISPCPVP VARcaIgGD ESLGGPSVFIFPPKPKDTLLIARTPEITCVVL DLGREDPEVQISWFVDGKEVHTAKTQPREQQF NSTYRVVSVLPIGHQDWLTGKEFKCRVNHIGL PSPIERTISKARGQAHQPSVYVLPPSPKELSS SDTVTLTCLIKDFFPPEIDVEWQSNGQPEPES KYHTTAPQLDEDGSYFLYSKLSVDKSRWQQGD TFTCAVMHEALHNHYTDLSLSHSPGKGGGGSG GGGHAEGTFTSDVSSYLEGQAAKEFIAWLVKG GGG 56 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8/Gluc-4G_IV_ GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG VAReqIgGD GPSVFIFPPNPKDTLMISRTPVVTCVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VTCLVKDFYPPDISVEWQSNRWPELEGKYSTT PAQLDGDGSYFLYSKLSLETSRWQQVESFICA
VMHEALHNHYTKTDISESLGKGGGGSGGGGHS QGTFTSDYSKYLDSRRAQDFVQWLMNTGGGG 57 HSQGTFTSDYSKYLDSRRAQDFVQWLMNTGAG Gluc/GLP1-3G_V_ GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG VAReqIgG2 GPSVFIFPPNPKDTLMISRTPVVTCVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VTCLVKDFYPPDISVEWQSNRWPELEGKYSTT PAQLDGDGSYFLYSKLSLETSRWQQVESFICA VMHEALHNHYTKTDISESLGKGGGGSGGGGHA EGTFTSDVSSYLEGQAAKEFIAWLVKGGGG 58 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8/Glu-4G_IV_ GGGGSGGGSGGGSGGGSSESKYGPPCPPCPAP huIgG4 EFLGGPSVFLFPPKPKDTLMISRTPEVICVVV DVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGL PSSIEKTISKAKGQPREPQVYTLPPSQEEMTK NQVSLTCLVKGFYPSDIAVEWESNGQPENNYK TTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFS CSVMHEALHNHYTQKSLSLSLGKGGGGSGGGG HSQGTFTSDYSKYLDSRRAQDFVQWLMNTGGG G 59 HSQGTFTSDYSKYLDSRRAQDFVQWLMNTGGG Gluc/GLP1-3G_V_huIgG4 GGSGGGSGGGSGGGSSESKYGPPCPPCPAPEF LGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV SQEDPEVQFNWYVDGVEVHNAKTKPREEQFNS TYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQ VSLTCLVKGFYPSDIAVEWESNGQPENNYKTT PPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCS VMHEALHNHYTQKSLSLSLGKGGGGSGGGGHA EGTFTSDVSSYLEGQAAKEFIAWLVKGGAGGG G 60 PVPEPLGGPSVLIFPPKPKDTLRITRTPEVIC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE IgG-A Fc QQFNGTYRVVSVLPIEHQDWLTGKEFKCRVNH C1q- IDLPSPIERTISKARGRAHKPSVYVLPPSPKE Protein A+ LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE I(21)T PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ Q(207)H QGDPFTCAVMHETLHNHYTDLSLSHSPGK 61 PGCGLLGGPSVFIFPPKPKDTLVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN C1q+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE Protein A+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP I(21)T ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 62 PVPESLGGPSVFIFPPKPKDTLRITRTPEITC Exemplary variant canine VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE IgG-D Fc QQFNSTYRVVSVLPIEHQDWLTGKEFKCRVNH C1q- IGLPSPIERTISKARGQAHQPSVYVLPPSPKE Protein A+ LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE I(21)T PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ Q(207)H QGDTFTCAVMHEALHNHYTDLSLSHSPGK 63 GGPSVFLFPPNPKDILMITRIPEVICVVVDVS Exemplary wild-type equine QENPDVKFNWYMDGVEVRTATTRPKEEQFNST IgG1 Fc YRVVSVLRIQHQDWLSGKEFKCKVNNQALPQP Protein A+ IERTITKTKGRSQEPQVYVLAPHPDESKKSKV C1q+ SVTCLVKDFYPPEINIEWQSNGQPELETKYST TQAQQDSDGSYFLYSKLSVDRNRWQQGTTFTC GVMHEALHNHYTQKNVSKNPGK 64 GGPSVFIFPPNPKDALMISRTPVVTCVVVNLS Exemplary wild-type equine DQYPDVQFSWYVDNTEVHSAITKQREAQFNST IgG2 Fc YRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQP Protein A- ISRAISRGKGPSRVPQVYVLPPHPDELAKSKV C1q- SVTCLVKDFYPPDISVEWQSNRWPELEGKYST TPAQLDGDGSYFLYSKLSLETSRWQQVESFTC AVMHEALHNHFTKTDISESLGK 65 GGPSVFIFPPKPKDVLMITRMPEVTCLVVDVS Exemplary wild-type equine HDSSDVLFTWYVDGTEVKTAKTMPNEEQNNST IgG3 Fc YRVVSVLRIQHQDWLNGKKFKCKVNNQALPAP Protein A+ VERTISKATGQTRVPQVYVLAPHPDELSKNKV C1q+ SVTCLVKDFYPPDITVEWQSNEHPEPEGKYRT TEAQKDSDGSYFLYSKLTVEKDRWQQGTTFTC VVMHEALHNHVMQKNISKNPGK 66 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary wild-type equine HDFPDVQFNWYVDGVETHTATTEPKQEQFNST IgG4 Fc YRVVSVLPIQHKDWLSGKEFKCKVNNKALPAP Protein A+ VERTISAPTGQPREPQVYVLAPHRDELSKNKV C1q+ SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST TPAQLDSDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 67 GGPSVFIFPPKPKDVLMISRKPEVTCVVVDLG Exemplary wild-type equine HDDPDVQFTWFVDGVETHTATTEPKEEQFNST IgG5 Fc YRVVSVLPIQHQDWLSGKEFKCSVTSKALPAP Protein- VERTISKAKGQLRVPQVYVLAPHPDELAKNTV C1q- SVTCLVKDFYPPEIDVEWQSNEHPEPEGKYST TPAQLNSDGSYFLYSKLSVETSRWKQGESFTC GVMHEAVENHYTQKNVSHSPGK 68 GRPSVFIFPPNPKDTLMISRTPEVTCVVVDVS Exemplary wild-type equine QENPDVKFNWYVDGVEAHTATTKAKEKQDNST IgG6 Fc YRVVSVLPIQHQDWRRGKEFKCKVNNRALPAP Protein A- VERTITKAKGELQDPQVYILAPHPDEVTKNTV C1q- SVTCLVKDFYPPDINVEWQSNEEPEPEVKYST TPAQLDGDGSYFLYSKLTVETDRWEQGESFTC VVMHEAIRHTYRQKSITNFPGK 69 VGPSVFIFPPKPKDVLMISRTPTVICVVVDVG Exemplary wild-type equine HDFPDVQFNWYVDGVETHTATTEPKQEQNNST IgG7 Fc YRVVSILAIQHKDWLSGKEFKCKVNNQALPAP Protein A+ VQKTISKPTGQPREPQVYVLAPHPDELSKNKV C1q+ SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST TPAQLDGDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 70 GGPSVFLFPPNPKDILMITRIPEVICVVVDVS Exemplary variant equine QENPDVKFNWYMDGVEVRTATTRPKEEQFNST IgG1 Fc YRVVSVLRIQHQDWLSGKEFKCSVNNQALPQP Protein A+ IERTITKTKGRSQEPQVYVLAPHPDESKKSKV C1q- SVTCLVKDFYPPEINIEWQSNGQPELETKYST K(87)S TQAQQDSDGSYFLYSKLSVDRNRWQQGTTFTC GVMHEALHNHYTQKNVSKNPGK 71 GGPSVFIFPPNPKDALMISRTPVVICVVVNLS Exemplary variant equine DQYPDVQFSWYVDNTEVHSAITKQREAQFNST IgG2 Fc YRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQP C1q- ISRAISRGKGPSRVPQVYVLPPHPDELAKSKV Protein A+ SVTCLVKDFYPPDISVEWQSNRWPELEGKYST F(203)Y TPAQLDGDGSYFLYSKLSLETSRWQQGESFTC AVMHEALHNHYTKTDISESLGK 72 GGPSVFIFPPNPKDTLMISRTPVVICVVVNLS Exemplary variant equine DQYPDVQFSWYVDNTEVHSAITKQREAQFNST IgG2 Fc YRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQP C1q- ISRAISRGKGPSRVPQVYVLPPHPDELAKSKV Protein A+ SVTCLVKDFYPPDISVEWQSNRWPELEGKYST A(15)T TPAQLDGDGSYFLYSKLSLETSRWQQVESFTC F(203)Y AVMHEALHNHYTKTDISESLGK 129 PPCVLSAEGVIPIPSVPKPQCPPYTHSKFLGG Exemplary wild-type equine PSVFIFPPNPKDALMISRTPVVTCVVVNLSDQ IgG2 Fc with hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A- VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP AQLDGDGSYFLYSKLSLETSRWQQVESFTCAV MHEALHNHFTKTDISESLGK 130 PPCVLSAEGVIPIPSVPKPPCPPYTHSKFLGG Exemplary variant equine PSVFIFPPNPKDALMISRTPVVTCVVVNLSDQ IgG2 Fc with modified hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A- VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV Q(20)P TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP AQLDGDGSYFLYSKLSLETSRWQQVESFTCAV MHEALHNHFTKTDISESLGK 131 PPSVLSAEGVIPIPSVPKPQCPPYTHSKFLGG Exemplary variant equine PSVFIFPPNPKDALMISRTPVVTCVVVNLSDQ IgG2 Fc with modified hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A- VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV C(3)S TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP AQLDGDGSYFLYSKLSLETSRWQQVESFTCAV MHEALHNHFTKTDISESLGK 132 PPSVLSAEGVIPIPSVPKPPCPPYTHSKFLGG Exemplary variant equine PSVFIFPPNPKDALMISRTPVVTCVVVNLSDQ IgG2 Fc with modified hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A- VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV C(3)S TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP Q(20)P AQLDGDGSYFLYSKLSLETSRWQQVESFICAV MHEALHNHFTKTDISESLGK 133 PPCVLSAEGVIPIPSVPKPQCPPYTHSKFLGG Exemplary variant equine PSVFIFPPNPKDTLMISRTPVVICVVVNLSDQ IgG2 Fc with hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A+ VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV A(45)T TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP F(233)Y AQLDGDGSYFLYSKLSLETSRWQQVESFICAV MHEALHNHYTKTDISESLGK 134 PPCVLSAEGVIPIPSVPKPPCPPYTHSKFLGG Exemplary variant equine PSVFIFPPNPKDTLMISRTPVVTCVVVNLSDQ IgG2 Fc with modified hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A+ VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV Q(20)P TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP A(45)T AQLDGDGSYFLYSKLSLETSRWQQVESFICAV F(233)Y MHEALHNHYTKTDISESLGK 135 PPSVLSAEGVIPIPSVPKPPCPPYTHSKFLGG Exemplary variant equine PSVFIFPPNPKDTLMISRTPVVTCVVVNLSDQ IgG2 Fc with modified hinge YPDVQFSWYVDNTEVHSAITKQREAQFNSTYR Protein A+ VVSVLPIQHQDWLSGKEFKCSVTNVGVPQPIS C1q- RAISRGKGPSRVPQVYVLPPHPDELAKSKVSV C(3)S TCLVKDFYPPDISVEWQSNRWPELEGKYSTTP Q(20)P AQLDGDGSYFLYSKLSLETSRWQQVESFICAV A(45)T MHEALHNHYTKTDISESLGK F(233)Y 73 GGPSVFIFPPKPKDVLMITRMPEVTCLVVDVS Exemplary variant equine HDSSDVLFTWYVDGTEVKTAKTMPNEEQNNST IgG3 Fc YRVVSVLRIQHQDWLNGKKFKCSVNNQALPAP Protein A+ VERTISKATGQTRVPQVYVLAPHPDELSKNKV C1q- SVTCLVKDFYPPDITVEWQSNEHPEPEGKYRT K(87)S TEAQKDSDGSYFLYSKLTVEKDRWQQGTTFTC VVMHEALHNHVMQKNISKNPGK 74 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary variant equine HDFPDVQFNWYVDGVETHTATTEPKQEQFNST IgG4 Fc YRVVSVLPIQHKDWLSGKEFKCSVNNKALPAP Protein A+ VERTISAPTGQPREPQVYVLAPHRDELSKNKV C1q- SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST K(87)S TPAQLDSDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 75 GGPSVFIFPPKPKDVLMISRKPEVTCVVVDLG Exemplary variant equine HDDPDVQFTWFVDGVETHTATTEPKEEQFNST IgG5 Fc YRVVSVLPIQHQDWLSGKEFKCSVTSKALPAP C1q- VERTISKAKGQLRVPQVYVLAPHPDELAKNTV Protein A+ SVTCLVKDFYPPEIDVEWQSNEHPEPEGKYST V(199)L TPAQLNSDGSYFLYSKLSVETSRWKQGESFTC E(200)H GVMHEALHNHYTQKNVSHSPGK 76 GRPSVFIFPPNPKDTLMISRTPEVTCVVVDVS Exemplary variant equine QENPDVKFNWYVDGVEAHTATTKAKEKQDNST IgG6 Fc YRVVSVLPIQHQDWRRGKEFKCKVNNRALPAP C1q- VERTITKAKGELQDPQVYILAPHPDEVTKNTV Protein A+ SVTCLVKDFYPPDINVEWQSNEEPEPEVKYST I(199)L TPAQLDGDGSYFLYSKLTVETDRWEQGESFTC R(200)H VVMHEALHNHYRQKSITNFPGK H(201)N T(202)H 77 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary variant equine HDFPDVQFNWYVDGVETHTATTEPKQEQNNST IgG7 Fc YRVVSILAIQHKDWLSGKEFKCSVNNQALPAP Protein A+ VQKTISKPTGQPREPQVYVLAPHPDELSKNKV C1q- SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST K(87)S TPAQLDGDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 78 PAPEMLGGPSVFIFPPKPKDTLLIARTPEVTC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCRVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q- LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP K(93)R
ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 79 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCRVNN Protein A- KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q- MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP K(93)R ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 80 RKTDHPPGPKTGEGPKCPPPEMLGGPSIFIFP Exemplary wild-type feline PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT IgG1a Fc WFVDNTQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEEKCKVNSKSLPSPIERTISKAK C1q+ GQPHEPQVYVLPPAQEELSENKVSVTCLIKSF HPPDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 81 RKTDHPPGPKTGEGPKCPPPEMLGGPSIFIFP Exemplary wild-type feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1b Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEEKCKVNSKSLPSPIERTISKDK C1q+ GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF YPSDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 136 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1a Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEEKCKVNSKSLPSPIERTISKAK C1q- GQPHEPQVYVLPPAQEELSENKVSVICLIKSF P(198)A HPPDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 82 RKTDHPPGPKTGEGPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgGla Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEEKCKVNSKSLPSPIERTISKAK C1q- GQPHEPQVYVLPPAQEELSENKVSVICLIKSF P(198)A HPPDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 137 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1b Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEEKCKVNSKSLPSPIERTISKDK C1q- GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF P(198)A YPSDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 83 RKTDHPPGPKTGEGPKCPPPEMLGGPSIFIFP Exemplary variant feline PKPKDTLSISRTPEVICLVVDLGPDDSDVQIT IgG1b Fc WFVDNIQVYTAKTSPREEQFNSTYRVVSVLPI Protein A+ LHQDWLKGKEEKCKVNSKSLPSPIERTISKDK C1q- GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF P(198)A YPSDIAVEWEITGQPEPENNYRTIPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 84 PGCGLLGGPSVFIFPPKPKDTLLIARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQEKCRVNN C1q- KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE K(93)R MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP Protein A+ ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR I(21)T GDTFICAVMHEALHNHYTQISLSHSPGK V(23)L T(24)I 85 HAEGTFTSDVSSYLEGQAAKEFIAWLVKGAG Wild-type GLP1 (7-37) 86 HSEGTFTSDVSSYLEGQAAKEFIAWLVKG GLP1-S8 (7-35) 87 HGEGTFTSDVSSYLEGQAAKEFIAWLVKG GLP1-G8 (7-35) 88 G 1G extension 89 GG 2G extension 90 GGG 3G extension 91 GGGG 4G extension 92 GGGGG 5G extension 93 GGGGGG 6G extension 94 GGGGGGG 7G extension 95 GGGGGGGG 8G extension 96 MAVLGLLFCLVTFPSCVLSHGEGIFTSDVSSY ssGLP1-G8_I_WTfeIgG2 LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS GGGSPKTASTIESKTGEGPKCPVPEIPGAPSV FIFPPKPKDTLSISRTPEVTCLVVDLGPDDSN VQITWFVDNTEMHTAKTRPREEQFNSTYRVVS VLPILHQDWLKGKEFKCKVNSKSLPSAMERTI SKAKGQPHEPQVYVLPPTQEELSENKVSVTCL IKGFHPPDIAVEWEITGQPEPENNYQTTPPQL DSDGTYFLYSRLSVDRSHWQRGNTYTCSVSHE ALHSHHTQKSLTQSPGK 97 MAVLGLLFCLVTFPSCVLSHGEGIFTSDVSSY ssGLP1-G8/GLP1-3G_III_ LEGQAAKEFIAWLVKGGAGGGGGSGGGSGGGS WTfeIgG2 GGGSPKTASTIESKTGEGPKCPVPEIPGAPSV FIFPPKPKDTLSISRTPEVTCLVVDLGPDDSN VQITWFVDNTEMHTAKTRPREEQFNSTYRVVS VLPILHQDWLKGKEFKCKVNSKSLPSAMERTI SKAKGQPHEPQVYVLPPTQEELSENKVSVTCL IKGFHPPDIAVEWEITGQPEPENNYQTTPPQL DSDGTYFLYSRLSVDRSHWQRGNTYTCSVSHE ALHSHHTQKSLTQSPGKGGGGSGGGGHAEGTF TSDVSSYLEGQAAKEFIAWLVKGGGG 98 HAEGTFTSDVSSYLEGQAAKEFIAWLVKGA Variant GLP1 (7-36) 99 HAEGTFTSDVSSYLEGQAAKEFIAWLVKG Variant GLP1 (7-35) 100 VPKPQCPPYTHSKFLGGPSVFIFPPNPKDALM Exemplary wild-type equine ISRTPVVTCVVVNLSDQYPDVQFSTNYVDNTEV Fc IgG2 (including equine HSAITKQREAQFNSTYRVVSVLPIQHQDWLSG IgG2 hinge) KEFKCSVTNVGVPQPISRAISRGKGPSRVPQV Protein- YVLPPHPDELAKSKVSVTCLVKDFYPPDISVE C1q- WQSNRWPELEGKYSTTPAQLDGDGSYFLYSKL SLETSRWQQVESFTCAVMHEALHNHFIKTDIS ESLGK 101 DMSKCPKCPAPELL Exemplary wild-type equine IgG1 hinge 102 VPKPQCPPYTHSKFL Exemplary wild-type equine IgG2 hinge 138 PPCVLSAEGVIPIPSVPKPQCPPYTHSKFL Exemplary wild-type equine IgG2 hinge 103 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8/GLP1-3G_III_ GGGGSGGGSGGGSGGGSDMSKCPKCPAPELLG VAReqIgG2 GPSVFIFPPNPKDTLMISRTPVVICVVVNLSD QYPDVQFSWYVDNTEVHSAITKQREAQFNSTY RVVSVLPIQHQDWLSGKEFKCSVTNVGVPQPI SRAISRGKGPSRVPQVYVLPPHPDELAKSKVS VTCLVKDFYPPDISVEWQSNRWPELEGKYSTT PAQLDGDGSYFLYSKLSLETSRWQQVESFTCA VMHEALHNHYTKTDISESLGKGGGGSGGGGHA EGTFTSDVSSYLEGQAAKEFIAWLVKGGGG 104 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGAG GLP1-G8_I_VAReqIgG2 GGGGSGGGSGGGSGGGSSVPKPQCPPYTHSKF LGGPSVFIFPPNPKDTLMISRTPVVTCVVVNL SDQYPDVQFSWYVDNTEVHSAITKQREAQFNS TYRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQ PISRAISRGKGPSRVPQVYVLPPHPDELAKSK VSVTCLVKDFYPPDISVEWQSNRWPELEGKYS TTPAQLDGDGSYFLYSKLSLETSRWQQVESFT CAVMHEALHNHYTKTDISESLGK 105 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8/GLP1-2G_III_ SGGGGSGGGGSGGGGSPKESTCKCISPCPVPE VARcaIgGD SLGGPSVFIFPPKPKDTLLIARTPEITCVVLD LGREDPEVQISWFVDGKEVHTAKTQPREQQFN STYRVVSVLPIGHQDWLTGKEFKCRVNHIGLP SPIERTISKARGQAHQPSVYVLPPSPKELSSS DTVTLTCLIKDFFPPEIDVEWQSNGQPEPESK YHTTAPQLDEDGSYFLYSKLSVDKSRWQQGDT FTCAVMHEALHNHYTDLSLSHSPGKGGGGSGG GGHAEGTFTSDVSSYLEGQAAKEFIAWLVKGG G 106 HGEGTFTSDVSSYLEGQAAKEFIAWLVKGGGG GLP1-G8/GLP1-2G_III_ SGGGGSGGGGSGGGGSPKESTSKCISPCPVPE VARcaIgGD SLGGPSVFIFPPKPKDTLLIARTPEITCVVLD LGREDPEVQISWFVDGKEVHTAKTQPREQQFN STYRVVSVLPIGHQDWLTGKEFKCRVNHIGLP SPIERTISKARGQAHQPSVYVLPPSPKELSSS DTVTLTCLIKDFFPPEIDVEWQSNGQPEPESK YHTTAPQLDEDGSYFLYSKLSVDKSRWQQGDT FTCAVMHEALHNHYTDLSLSHSPGKGGGGSGG GGHAEGTFTSDVSSYLEGQAAKEFIAWLVKGG G 139 PAPEPLGGPSVFIFPPKPKDTLLTARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR M(5)P GDTFICAVMHEALHNHYTQESLSHSPGK 140 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDREDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR P(39)R GDTFICAVMHEALHNHYTQESLSHSPGK 141 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLGPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR D(38)G GDTFICAVMHEALHNHYTQESLSHSPGK 142 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ IALPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR K(97)I GDTFICAVMHEALHNHYTQESLSHSPGK 143 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ KGLPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR A(98)G GDTFICAVMHEALHNHYTQESLSHSPGK 144 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLGPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ IGLPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR D(38)G GDTFICAVMHEALHNHYTQESLSHSPGK K(97)I A(98)G 145 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLGPEDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCRVNN Protein A+ IGLPSPIERTISKARGQAHQPSVYVLPPSREE C1q- LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR D(38)G GDTFICAVMHEALHNHYTQESLSHSPGK K(93)R K(97)I A(98)G 146 PAPEPLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDREDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCKVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q+ LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR M(5)P
GDTFICAVMHEALHNHYTQESLSHSPGK P(39)R 147 PAPEPLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary variant canine VVVDLDREDPEVQISWFVDGKQMQTAKTQPRE IgG-B Fc EQFNGTYRVVSVLPIGHQDWLKGKQFTCRVNN Protein A+ KALPSPIERTISKARGQAHQPSVYVLPPSREE C1q- LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP CD16- ESKYRTIPPQLDEDGSYFLYSKLSVDKSRWQR M(5)P GDTFICAVMHEALHNHYTQESLSHSPGK P(39)R K(93)R 148 PGCGPLGGPSVFIFPPKPKDILVTARTPTVIC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR L(5)P GDTFICAVMHEALHNHYTQISLSHSPGK 149 PGCGLLGGPSVFIFPPKPKDILVTARTPTVIC Exemplary variant canine VVVDLDRENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR P(39)R GDTFICAVMHEALHNHYTQISLSHSPGK 150 PGCGLLGGPSVFIFPPKPKDILVTARTPTVIC Exemplary variant canine VVVDLGPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR D(38)G GDTFICAVMHEALHNHYTQISLSHSPGK 151 PGCGLLGGPSVFIFPPKPKDILVTARTPTVIC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ IALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR K(97)I GDTFICAVMHEALHNHYTQISLSHSPGK 152 PGCGLLGGPSVFIFPPKPKDILVTARTPTVIC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ KGLPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR A(98)G GDTFICAVMHEALHNHYTQISLSHSPGK 153 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCRVNN Protein A+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q- MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16+ ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR K(93)R GDTFICAVMHEALHNHYTQISLSHSPGK 154 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLGPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ IGLPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR D(38)G GDTFICAVMHEALHNHYTQISLSHSPGK K(97)I A(98)G 155 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLGPENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCRVNN Protein A+ IGLPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q- MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR D(38)G GDTFICAVMHEALHNHYTQISLSHSPGK K(93)R K(97)I A(98)G 156 PGCGPLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDRENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCKVNN Protein A+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q+ MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR L(5)P GDTFICAVMHEALHNHYTQISLSHSPGK P(39)R 157 PGCGPLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary variant canine VVVDLDRENPEVQISWFVDSKQVQTANTQPRE IgG-C Fc EQSNGTYRVVSVLPIGHQDWLSGKQFKCRVNN Protein A+ KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE C1q- MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP CD16- ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR M(5)P GDTFICAVMHEALHNHYTQISLSHSPGK P(39)R K(93)R 158 PVPEPLGGPSVLIFPPKPKDILRITRTPEVIC Exemplary aglycosyl variant VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE canine IgG-A Fc QQFX.sub.1GTYRVVSVLPIEHQDWLIGKEFKCRVNH N(68)X.sub.1 IDLPSPIERTISKARGRAHKPSVYVLPPSPKE X.sub.1 = any amino acid except N LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 159 PVPEPLGGPSVLIFPPKPKDILRITRTPEVIC Exemplary aglycosyl variant VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE canine IgG-A Fc QQFNPTYRVVSVLPIEHQDWLTGKEFKCRVNH G(69)13 IDLPSPIERTISKARGRAHKPSVYVLPPSPKE LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 160 PVPEPLGGPSVLIFPPKPKDILRITRTPEVIC Exemplary aglycosyl variant VVLDLGREDPEVQISWFVDGKEVHTAKTQSRE canine IgG-A Fc QQFNGX.sub.2YRVVSVLPIEHQDWLTGKEFKCRVNH T(70)X.sub.2 IDLPSPIERTISKARGRAHKPSVYVLPPSPKE X.sub.2 = any amino acid except T LSSSDTVSITCLIKDFYPPDIDVEWQSNGQQE or S PERKHRMTPPQLDEDGSYFLYSKLSVDKSRWQ QGDPFTCAVMHETLQNHYTDLSLSHSPGK 161 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary aglycosyl variant VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE canine IgG-B Fc EQFX.sub.1GTYRVVSVLPIGHQDWLKGKQFTCKVNN N(68)X.sub.1 KALPSPIERTISKARGQAHQPSVYVLPPSREE X.sub.1 = any amino acid except N LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 162 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary aglycosyl variant VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE canine IgG-B Fc EQFNPTYRVVSVLPIGHQDWLKGKQFTCKVNN G(69)13 KALPSPIERTISKARGQAHQPSVYVLPPSREE LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 163 PAPEMLGGPSVFIFPPKPKDILLIARTPEVIC Exemplary aglycosyl variant VVVDLDPEDPEVQISWFVDGKQMQTAKTQPRE canine IgG-B Fc EQFNGX.sub.2YRVVSVLPIGHQDWLKGKQFTCKVNN T(70)X.sub.2 KALPSPIERTISKARGQAHQPSVYVLPPSREE X.sub.2 = any amino acid except T LSKNTVSLTCLIKDFFPPDIDVEWQSNGQQEP or S ESKYRTTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQESLSHSPGK 164 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary aglycosyl variant VVVDLDPENPEVQISWFVDSKQVQTANTQPRE canine IgG-C Fc EQSX.sub.1GTYRVVSVLPIGHQDWLSGKQFKCKVNN N(68)X.sub.1 KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE X.sub.1 = any amino acid except N MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 165 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary aglycosyl variant VVVDLDPENPEVQISWFVDSKQVQTANTQPRE canine IgG-C Fc EQSNPTYRVVSVLPIGHQDWLSGKQFKCKVNN G(69)13 KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 166 PGCGLLGGPSVFIFPPKPKDILVTARTPTVTC Exemplary aglycosyl variant VVVDLDPENPEVQISWFVDSKQVQTANTQPRE canine IgG-C Fc EQSNGX.sub.2YRVVSVLPIGHQDWLSGKQFKCKVNN T(70)X.sub.2 KALPSPIEEIISKTPGQAHQPNVYVLPPSRDE X.sub.2 = any amino acid except T MSKNTVTLTCLVKDFFPPEIDVEWQSNGQQEP or S ESKYRMTPPQLDEDGSYFLYSKLSVDKSRWQR GDTFICAVMHEALHNHYTQISLSHSPGK 167 PVPESLGGPSVFIFPPKPKDILRITRTPEITC Exemplary aglycosyl variant VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE canine IgG-D Fc QQFX.sub.1STYRVVSVLPIEHQDWLTGKEFKCRVNH N(68)X.sub.1 IGLPSPIERTISKARGQAHQPSVYVLPPSPKE X.sub.1 = any amino acid except N LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 168 PVPESLGGPSVFIFPPKPKDILRITRTPEITC Exemplary aglycosyl variant VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE canine IgG-D Fc QQFNPTYRVVSVLPIEHQDWLTGKEFKCRVNH S(69)P IGLPSPIERTISKARGQAHQPSVYVLPPSPKE LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 169 PVPESLGGPSVFIFPPKPKDILRITRTPEITC Exemplary aglycosyl variant VVLDLGREDPEVQISWFVDGKEVHTAKTQPRE canine IgG-D Fc QQFNSX.sub.2YRVVSVLPIEHQDWLTGKEFKCRVNH T(70)X.sub.2 IGLPSPIERTISKARGQAHQPSVYVLPPSPKE X.sub.2 = any amino acid except T LSSSDTVTLTCLIKDFFPPEIDVEWQSNGQPE or S PESKYHTTAPQLDEDGSYFLYSKLSVDKSRWQ QGDTFTCAVMHEALQNHYTDLSLSHSPGK 170 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT feline IgG1a Fc WFVDNTQVYTAKTSPREEQFX.sub.1STYRVVSVLPI N(85)X.sub.1 LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK X.sub.1 = any amino acid except N GQPHEPQVYVLPPAQEELSENKVSVTCLIKSF HPPDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 171 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT feline IgG1a Fc WFVDNTQVYTAKTSPREEQFNPTYRVVSVLPI S(86)P LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK GQPHEPQVYVLPPAQEELSENKVSVTCLIKSF HPPDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 172 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT feline IgG1a Fc WFVDNTQVYTAKTSPREEQFNSX.sub.2YRVVSVLPI T(87)X.sub.2 LHQDWLKGKEFKCKVNSKSLPSPIERTISKAK X.sub.2 = any amino acid except T GQPHEPQVYVLPPAQEELSENKVSVTCLIKSF or S HPPDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFVYSKLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 173 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT feline IgG1b Fc WFVDNTQVYTAKTSPREEQFX.sub.1STYRVVSVLPI N(85)X.sub.1 LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK X.sub.1 = any amino acid except N GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF YPSDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 174 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT feline IgG1b Fc WFVDNTQVYTAKTSPREEQFNPTYRVVSVLPI S(86)P LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF YPSDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 175 RKTDHPPGPKPCDCPKCPPPEMLGGPSIFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSDVQIT feline IgG1b Fc WFVDNTQVYTAKTSPREEQENSX.sub.2YRVVSVLPI T(87)X.sub.2 LHQDWLKGKEFKCKVNSKSLPSPIERTISKDK X.sub.2 = any amino acid except T GQPHEPQVYVLPPAQEELSENKVSVTCLIEGF or S YPSDIAVEWEITGQPEPENNYRTTPPQLDSDG TYFLYSRLSVDRSRWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 176 PKTASTIESKTGEGPKCPVPEIPGAPSVFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSNVQIT feline IgG2 Fc WFVDNTEMHTAKTRPREEQEX.sub.1STYRVVSVLPI N(85)X.sub.1 LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK X.sub.1 = any amino acid except N GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG
TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 177 PKTASTIESKTGEGPKCPVPEIPGAPSVFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSNVQIT feline IgG2 Fc WFVDNTEMHTAKTRPREEQFNPTYRVVSVLPI S(86)P LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 178 PKTASTIESKTGEGPKCPVPEIPGAPSVFIFP Exemplary aglycosyl variant PKPKDTLSISRTPEVTCLVVDLGPDDSNVQIT feline IgG2 Fc WFVDNTEMHTAKTRPREEQENSX.sub.2YRVVSVLPI T(87)X.sub.2 LHQDWLKGKEFKCKVNSKSLPSAMERTISKAK X.sub.2 = any amino acid except T GQPHEPQVYVLPPTQEELSENKVSVTCLIKGF or S HPPDIAVEWEITGQPEPENNYQTTPPQLDSDG TYFLYSRLSVDRSHWQRGNTYTCSVSHEALHS HHTQKSLTQSPGK 179 GGPSVFLEPPNPKDILMITRIPEVICVVVDVS Exemplary aglycosyl variant QENPDVKFNWYMDGVEVRTATTRPKEEQFX.sub.1ST equine IgG1 Fc YRVVSVLRIQHQDWLSGKEFKCKVNNQALPQP N(62)X.sub.1 IERTITKTKGRSQEPQVYVLAPHPDESKKSKV X.sub.1 = any amino acid except N SVTCLVKDFYPPEINIEWQSNGQPELETKYST TQAQQDSDGSYFLYSKLSVDRNRWQQGTTFTC GVMHEALHNHYTQKNVSKNPGK 180 GGPSVFLEPPNPKDILMITRIPEVICVVVDVS Exemplary aglycosyl variant QENPDVKFNWYMDGVEVRTATTRPKEEQFNPT equine IgG1 Fc YRVVSVLRIQHQDWLSGKEFKCKVNNQALPQP S(63)P IERTITKTKGRSQEPQVYVLAPHPDESKKSKV SVTCLVKDFYPPEINIEWQSNGQPELETKYST TQAQQDSDGSYFLYSKLSVDRNRWQQGTTFTC GVMHEALHNHYTQKNVSKNPGK 181 GGPSVFLFPPNPKDILMITRIPEVICVVVDVS Exemplary aglycosyl variant QENPDVKFNWYMDGVEVRTATTRPKEEQFNSX.sub.2 equine IgG1 Fc YRVVSVLRIQHQDWLSGKEFKCKVNNQALPQP T(64)X.sub.2 IERTITKTKGRSQEPQVYVLAPHPDESKKSKV X.sub.2 = any amino acid except T SVTCLVKDFYPPEINIEWQSNGQPELETKYST or S TQAQQDSDGSYFLYSKLSVDRNRWQQGTTFTC GVMHEALHNHYTQKNVSKNPGK 182 GGPSVFIFPPNPKDALMISRTPVVTCVVVNLS Exemplary aglycosyl variant DQYPDVQFSWYVDNTEVHSAITKQREAQFX.sub.1ST equine IgG2 Fc YRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQP N(62)X.sub.1 ISRAISRGKGPSRVPQVYVLPPHPDELAKSKV X.sub.1 = any amino acid except N SVTCLVKDFYPPDISVEWQSNRWPELEGKYST TPAQLDGDGSYFLYSKLSLETSRWQQVESFTC AVMHEALHNHFTKTDISESLGK 183 GGPSVFIFPPNPKDALMISRTPVVTCVVVNLS Exemplary aglycosyl variant DQYPDVQFSWYVDNTEVHSAITKQREAQFNPT equine IgG2 Fc YRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQP S(63)P ISRAISRGKGPSRVPQVYVLPPHPDELAKSKV SVTCLVKDFYPPDISVEWQSNRWPELEGKYST TPAQLDGDGSYFLYSKLSLETSRWQQVESFTC AVMHEALHNHFTKTDISESLGK 184 GGPSVFIFPPNPKDALMISRTPVVTCVVVNLS Exemplary aglycosyl variant DQYPDVQFSWYVDNTEVHSAITKQREAQFNSX.sub.2 equine IgG2 Fc YRVVSVLPIQHQDWLSGKEFKCSVTNVGVPQP T(64)X.sub.2 ISRAISRGKGPSRVPQVYVLPPHPDELAKSKV X.sub.2 = any amino acid except T SVTCLVKDFYPPDISVEWQSNRWPELEGKYST or S TPAQLDGDGSYFLYSKLSLETSRWQQVESFTC AVMHEALHNHFTKTDISESLGK 185 GGPSVFIFPPKPKDVLMITRMPEVTCLVVDVS Exemplary aglycosyl variant HDSSDVLFTWYVDGTEVKTAKTMPNEEQNX.sub.1ST equine IgG3 Fc YRVVSVLRIQHQDWLNGKKFKCKVNNQALPAP N(62)X.sub.1 VERTISKATGQTRVPQVYVLAPHPDELSKNKV X.sub.1 = any amino acid except N SVTCLVKDFYPPDITVEWQSNEHPEPEGKYRT TEAQKDSDGSYFLYSKLTVEKDRWQQGTTFTC VVMHEALHNHVMQKNISKNPGK 186 GGPSVFIFPPKPKDVLMITRMPEVTCLVVDVS Exemplary aglycosyl variant HDSSDVLFTWYVDGTEVKTAKTMPNEEQNNPT equine IgG3 Fc YRVVSVLRIQHQDWLNGKKFKCKVNNQALPAP S(63)P VERTISKATGQTRVPQVYVLAPHPDELSKNKV SVTCLVKDFYPPDITVEWQSNEHPEPEGKYRT TEAQKDSDGSYFLYSKLTVEKDRWQQGTTFTC VVMHEALHNHVMQKNISKNPGK 187 GGPSVFIFPPKPKDVLMITRMPEVTCLVVDVS Exemplary aglycosyl variant HDSSDVLFTWYVDGTEVKTAKTMPNEEQNNSX.sub.2 equine IgG3 Fc YRVVSVLRIQHQDWLNGKKFKCKVNNQALPAP T(64)X.sub.2 VERTISKATGQTRVPQVYVLAPHPDELSKNKV X.sub.2 = any amino acid except T SVTCLVKDFYPPDITVEWQSNEHPEPEGKYRT or S TEAQKDSDGSYFLYSKLTVEKDRWQQGTTFTC VVMHEALHNHVMQKNISKNPGK 188 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary aglycosyl variant HDFPDVQFNWYVDGVETHTATTEPKQEQFX.sub.1ST equine IgG4 Fc YRVVSVLPIQHKDWLSGKEFKCKVNNKALPAP N(62)X.sub.1 VERTISAPTGQPREPQVYVLAPHRDELSKNKV X.sub.1 = any amino acid except N SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST IPAQLDSDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 189 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary aglycosyl variant HDFPDVQFNWYVDGVETHTATTEPKQEQFNPT equine IgG4 Fc YRVVSVLPIQHKDWLSGKEFKCKVNNKALPAP S(63)P VERTISAPTGQPREPQVYVLAPHRDELSKNKV SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST TPAQLDSDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 190 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary aglycosyl variant HDFPDVQFNWYVDGVETHTATTEPKQEQFNSX.sub.2 equine IgG4 Fc YRVVSVLPIQHKDWLSGKEFKCKVNNKALPAP T(64)X.sub.2 VERTISAPTGQPREPQVYVLAPHRDELSKNKV X.sub.2 = any amino acid except T SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST or S TPAQLDSDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 191 GGPSVFIFPPKPKDVLMISRKPEVTCVVVDLG Exemplary aglycosyl variant HDDPDVQFTWFVDGVETHTATTEPKEEQFX.sub.1PT equine IgG5 Fc YRVVSVLPIQHQDWLSGKEFKCSVTSKALPAP N(62)X.sub.1 VERTISKAKGQLRVPQVYVLAPHPDELAKNTV X.sub.1 = any amino acid except N SVTCLVKDFYPPEIDVEWQSNEHPEPEGKYST TPAQLNSDGSYFLYSKLSVETSRWKQGESFTC GVMHEAVENHYTQKNVSHSPGK 192 GGPSVFIFPPKPKDVLMISRKPEVTCVVVDLG Exemplary aglycosyl variant HDDPDVQFTWFVDGVETHTATTEPKEEQFNPT equine IgG5 Fc YRVVSVLPIQHQDWLSGKEFKCSVTSKALPAP S(63)P VERTISKAKGQLRVPQVYVLAPHPDELAKNTV SVTCLVKDFYPPEIDVEWQSNEHPEPEGKYST TPAQLNSDGSYFLYSKLSVETSRWKQGESFTC GVMHEAVENHYTQKNVSHSPGK 193 GGPSVFIFPPKPKDVLMISRKPEVTCVVVDLG Exemplary aglycosyl variant HDDPDVQFTWFVDGVETHTATTEPKEEQFNSX.sub.2 equine IgG5 Fc YRVVSVLPIQHQDWLSGKEFKCSVTSKALPAP T(64)X.sub.2 VERTISKAKGQLRVPQVYVLAPHPDELAKNTV X.sub.2 = any amino acid except T SVTCLVKDFYPPEIDVEWQSNEHPEPEGKYST or S TPAQLNSDGSYFLYSKLSVETSRWKQGESFTC GVMHEAVENHYTQKNVSHSPGK 194 GRPSVFIFPPNPKDTLMISRTPEVTCVVVDVS Exemplary aglycosyl variant QENPDVKFNWYVDGVEAHTATTKAKEKQDX.sub.1ST equine IgG6 Fc YRVVSVLPIQHQDWRRGKEFKCKVNNRALPAP N(62)X.sub.1 VERTITKAKGELQDPQVYILAPHPDEVTKNTV X.sub.1 = any amino acid except N SVTCLVKDFYPPDINVEWQSNEEPEPEVKYST TPAQLDGDGSYFLYSKLTVETDRWEQGESFTC VVMHEAIRHTYRQKSITNFPGK 195 GRPSVFIFPPNPKDTLMISRTPEVTCVVVDVS Exemplary aglycosyl variant QENPDVKFNWYVDGVEAHTATTKAKEKQDNPT equine IgG6 Fc YRVVSVLPIQHQDWRRGKEFKCKVNNRALPAP S(63)P VERTITKAKGELQDPQVYILAPHPDEVTKNTV SVTCLVKDFYPPDINVEWQSNEEPEPEVKYST IPAQLDGDGSYFLYSKLTVETDRWEQGESFTC VVMHEAIRHTYRQKSITNFPGK 196 GRPSVFIFPPNPKDTLMISRTPEVTCVVVDVS Exemplary aglycosyl variant QENPDVKFNWYVDGVEAHTATTKAKEKQDNSX.sub.2 equine IgG6 Fc YRVVSVLPIQHQDWRRGKEFKCKVNNRALPAP T(64)X.sub.2 VERTITKAKGELQDPQVYILAPHPDEVTKNTV X.sub.2 = any amino acid except T SVTCLVKDFYPPDINVEWQSNEEPEPEVKYST or S TPAQLDGDGSYFLYSKLTVETDRWEQGESFTC VVMHEAIRHTYRQKSITNFPGK 197 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary aglycosyl variant HDFPDVQFNWYVDGVETHTATTEPKQEQNX.sub.1ST equine IgG7 Fc YRVVSILAIQHKDWLSGKEFKCKVNNQALPAP N(62)X.sub.1 VQKTISKPTGQPREPQVYVLAPHPDELSKNKV X.sub.1 = any amino acid except N SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST TPAQLDGDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 198 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary aglycosyl variant HDFPDVQFNWYVDGVETHTATTEPKQEQNNPT equine IgG7 Fc YRVVSILAIQHKDWLSGKEFKCKVNNQALPAP S(63)P VQKTISKPTGQPREPQVYVLAPHPDELSKNKV SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST TPAQLDGDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK 199 VGPSVFIFPPKPKDVLMISRTPTVTCVVVDVG Exemplary aglycosyl variant HDFPDVQFNWYVDGVETHTATTEPKQEQNNSX.sub.2 equine IgG7 Fc YRVVSILAIQHKDWLSGKEFKCKVNNQALPAP T(64)X.sub.2 VQKTISKPTGQPREPQVYVLAPHPDELSKNKV X.sub.2 = any amino acid except T SVTCLVKDFYPPDIDIEWKSNGQPEPETKYST or S TPAQLDGDGSYFLYSKLTVETNRWQQGTTFTC AVMHEALHNHYTEKSVSKSPGK
DESCRIPTION OF THE EMBODIMENTS
[0262] Variant IgG Fc polypeptides from companion animals, such as canine, equine, and feline, are described. In some embodiments, the variant igG Fc polypeptides have increased binding to Protein A, decreased binding to C1q, decreased binding to CD16, increased stability, increased recombinant production, increased hinge disulfide formation, and/or form heterodimeric polypeptides. In some embodiments, antibodies, antibody fragments, or fusion proteins comprise a variant IgG Fc polypeptide. Methods of producing or purifying variant IgG Fc polypeptides and methods of administering variant IgG Fc polypeptides to companion animals are also provided assay.
[0263] Also provided are various embodiments relating to contiguous polypeptides and heterodimeric polypeptides comprising one or more variant GLP1 polypeptide(s) having improved serum half-life. In some embodiments, the contiguous polypeptides or heterodimeric polypeptides comprise a GLP1 polypeptide and a glucagon polypeptide as a dual GLP1 receptor and glucagon receptor agonist. In some embodiments, such polypeptides may be used to treat, for example, diabetes, obesity, or related indications, in companion animals, such as canines, felines, and equines.
[0264] For the convenience of the reader, the following definitions of terms used herein are provided.
[0265] As used herein, numerical terms such as K.sub.D are calculated based upon scientific measurements and, thus, are subject to appropriate measurement error. In some instances, a numerical term may include numerical values that are rounded to the nearest significant figure.
[0266] As used herein, "a" or "an" means "at least one" or "one or more" unless otherwise specified. As used herein, the term "or" means "and/or" unless specified otherwise. In the context of a multiple dependent claim, the use of "or" when referring back to other claims refers to those claims in the alternative only.
Exemplary Variant IgG Fc Polypeptides
[0267] Novel variant IgG Fc polypeptides are provided, for example, variant IgG Fc polypeptides for increased binding to Protein A, for decreased binding to C1q, for decreased binding to CD16, for increased stability, for increased recombinant production, for increased hinge disulfide formation, and/or for forming heterodimeric proteins assay.
[0268] "Amino acid sequence," means a sequence of amino acids residues in a peptide or protein. The terms "polypeptide" and "protein" are used interchangeably to refer to a polymer of amino acid residues, and are not limited to a minimum length. Such polymers of amino acid residues may contain natural or unnatural amino acid residues, and include, but are not limited to, peptides, oligopeptides, dimers, trimers, and multimers of amino acid residues. Both full-length proteins and fragments thereof are encompassed by the definition. The terms also include post-expression modifications of the polypeptide, for example, glycosylation, sialylation, acetylation, phosphorylation, and the like. Furthermore, for purposes of the present disclosure, a "polypeptide" refers to a protein which includes modifications, such as deletions, additions, and substitutions (generally conservative in nature), to the native sequence, as long as the protein maintains the desired activity. These modifications may be deliberate, as through site-directed mutagenesis, or may be accidental, such as through mutations of hosts which produce the proteins or errors due to PCR amplification.
[0269] A "fragment crystallizable polypeptide" or "Fc polypeptide" is the portion of an antibody molecule that interacts with effector molecules and cells. It comprises the C-terminal portions of the immunoglobulin heavy chains. As used herein, an Fc polypeptide includes fragments of the Fc domain having one or more biological activities of an entire Fc polypeptide. In some embodiments, a biological activity of an Fc polypeptide is the ability to bind FcRn. In some embodiments, a biological activity of an Fc polypeptide is the ability to bind C1q. In some embodiments, a biological activity of an Fc polypeptide is the ability to bind CD16. In some embodiments, a biological activity of an Fc polypeptide is the ability to bind protein A. An "effector function" of the Fc polypeptide is an action or activity performed in whole or in part by any antibody in response to a stimulus and may include complement fixation and/or ADCC (antibody-dependent cellular cytotoxicity) induction.
[0270] "IgX Fc" refers to an Fc polypeptide derived from a particular antibody isotype (e.g., IgG, IgA, IgD, IgE, IgM, etc.), where "X" denotes the antibody isotype. Thus, "IgG Fc" denotes that the Fc polypeptide is derived from a y chain, "IgA Fc" denotes that the Fc polypeptide is derived from an a chain, "IgD Fc" denotes that the Fc polypeptide is derived from a 6 chain, "IgE Fc" denotes that the Fc polypeptide is derived from a c chain, "IgM Fc" denotes that the Fc polypeptide is derived from a .mu. chain, etc. In some embodiments, the IgG Fc polypeptide comprises the hinge, CH2, and CH3, but does not comprise CH1 or CL. In some embodiments, the IgG Fc polypeptide comprises CH2 and CH3, but does not comprise CH1, the hinge, or CL. In some embodiments, the IgG Fc polypeptide comprises CH1, hinge, CH2, CH3, with or without CL. "IgX-N Fc" or "IgGXN Fc" denotes that the Fc polypeptide is derived from a particular subclass of antibody isotype (such as canine IgG subclass IgG-A, IgG-B, IgG-C, or IgG-D; feline IgG subclass IgG1a, IgG1b, or IgG2; or equine IgG subclass IgG1, IgG2, IgG3, IgG4, IgG5, IgG6, or IgG7, etc.), where "N" denotes the subclass.
[0271] "Hinge" refers to any portion of an Fc polypeptide or variant Fc polypeptide that is proline-rich and comprises at least one cysteine residue located between CH1 and CH2 of a full-length heavy chain constant region.
[0272] In some embodiments, a hinge is capable of forming a disulfide linkage within the same hinge region, within the same Fc polypeptide, with a hinge region of a separate Fc polypeptide, or with a separate Fc polypeptide. In some embodiments, a hinge comprises at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten proline residues.
[0273] The term "companion animal species" refers to an animal suitable to be a companion to humans. In some embodiments, a companion animal species is a canine (or dog), a feline (or cat), or an equine (or horse). In some embodiments, a companion animal species is a small mammal, such as a canine, feline, dog, cat, rabbit, ferret, guinea pig, rodent, etc. In some embodiments, a companion animal species is a farm animal, such as a horse, cow, pig, etc.
[0274] In some embodiments, an IgX Fc polypeptide or an IgX-N Fc polypeptide is derived from a companion animal, such as a dog, a cat, or a horse. In some embodiments, IgG Fc polypeptides are isolated from canine y heavy chains, such as IgG-A, IgG-B, IgG-C, or IgG-D. In some instances, IgG Fc polypeptides are isolated from feline y heavy chains, such as IgG1a, IgG1b, or IgG2. In other instances, IgG Fc polypeptides are isolated from equine y heavy chains, such as IgG1, IgG2, IgG3, IgG4, IgG5, IgG6, or IgG7.
[0275] The terms "IgX Fc" and "IgX Fc polypeptide" include wild-type IgX Fc polypeptides and variant IgX Fc polypeptides, unless indicated otherwise.
[0276] "Wild-type" refers to a non-mutated version of a polypeptide that occurs in nature, or a fragment thereof. A wild-type polypeptide may be produced recombinantly.
[0277] In some embodiments, a wild-type IgG Fc polypeptide comprises the amino acid sequence of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 16, SEQ ID NO: 63, SEQ ID NO: 64, SEQ ID NO: 65, SEQ ID NO: 66, SEQ ID NO: 67, SEQ ID NO: 68, SEQ ID NO: 69, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 100, SEQ ID NO: 107, SEQ ID NO: 108, SEQ ID NO: 117, SEQ ID NO: 118.
[0278] A "variant" is a polypeptide that differs from a reference polypeptide by single or multiple non-native amino acid substitutions, deletions, and/or additions. In some embodiments, a variant retains at least one biological activity of the reference polypeptide. In some embodiments, a variant (e.g., a variant canine IgG-A Fc, a variant canine IgG-C Fc, a variant canine IgG-D Fc, variant equine IgG2 Fc, variant equine IgG5 Fc, or variant equine IgG6 Fc) has an activity that the reference polypeptide substantially lacks. For example, in some embodiments, a variant canine IgG-A Fc, a variant canine IgG-C Fc, a variant canine IgG-D Fc, variant equine IgG2 Fc, variant equine IgG5 Fc, or variant equine IgG6 Fc binds Protein A.
[0279] As used herein, "percent (%) amino acid sequence identity" and "homology" with respect to a nucleic acid molecule or polypeptide sequence are defined as the percentage of nucleotide or amino acid residues in a reference sequence that are identical with the nucleotide or amino acid residues in the specific nucleic acid molecule or polypeptide sequence, after aligning the sequences and introducing gaps, if necessary to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Alignment for purposes of determining percent sequence identity can be achieved in various ways that are within the skill in the art, for instance, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or MEGALINE.TM. (DNASTAR) software. Those skilled in the art can determine appropriate parameters for measuring alignment, including any algorithms needed to achieve maximal alignment over the full length of sequences being compared.
[0280] In some embodiments, a variant has at least about 50% sequence identity with the reference nucleic acid molecule or polypeptide after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity, and not considering any conservative substitutions as part of the sequence identity. Such variants include, for instance, polypeptides wherein one or more amino acid residues are added, deleted, at the N- or C-terminus of the polypeptide. In some embodiments, a variant has at least about 50% sequence identity, at least about 60% sequence identity, at least about 65% sequence identity, at least about 70% sequence identity, at least about 75% sequence identity, at least about 80% sequence identity, at least about 85% sequence identity, at least about 90% sequence identity, at least about 95% sequence identity, at least about 97% sequence identity, at least about 98% sequence identity, or at least about 99% sequence identity with the sequence of the reference nucleic acid or polypeptide.
[0281] A "point mutation" is a mutation that involves a single amino acid residue. The mutation may be the loss of an amino acid, substitution of one amino acid residue for another, or the insertion of an additional amino acid residue.
[0282] An "amino acid substitution" refers to the replacement of one amino acid in a polypeptide with another amino acid. In some embodiments, an amino acid substitution is a conservative substitution. Nonlimiting exemplary conservative amino acid substitutions are shown in Table 2. Amino acid substitutions may be introduced into a molecule of interest and the products screened for a desired activity, for example, retained/improved antigen binding, decreased immunogenicity, or improved ADCC or CDC or enhanced pharmacokinetics.
TABLE-US-00002 TABLE 2 Original Residue Exemplary Substitutions Ala (A) Val; Leu; Ile Arg (R) Lys; Gln; Asn Asn (N) Gln; His; Asp; Lys; Arg Asp (D) Glu; Asn Cys (C) Ser; Ala Gln (Q) Asn; Glu Glu (E) Asp; Gln Gly (G) Ala His (H) Asn; Gln; Lys; Arg Ile (I) Leu; Val; Met; Ala; Phe; Norleucine Leu (L) Norleucine; Ile; Val; Met; Ala; Phe Lys (K) Arg; Gln; Asn Met (M) Leu; Phe; Ile Phe (F) Trp; Leu; Val; Ile; Ala; Tyr Pro (P) Ala Ser (S) Thr Thr (T) Val; Ser Trp (W) Tyr; Phe Tyr (Y) Trp; Phe; Thr; Ser Val (V) Ile; Leu; Met; Phe; Ala; Norleucine
[0283] Amino acids may be grouped according to common side-chain properties:
[0284] (1) hydrophobic: Norleucine, Met, Ala, Val, Leu, Ile;
[0285] (2) neutral hydrophilic: Cys, Ser, Thr, Asn, Gln;
[0286] (3) acidic: Asp, Glu;
[0287] (4) basic: His, Lys, Arg;
[0288] (5) residues that influence chain orientation: Gly, Pro;
[0289] (6) aromatic: Trp, Tyr, Phe.
[0290] Non-conservative substitutions entail exchanging a member of one of these classes with another class.
[0291] A "variant IgG Fc" as used herein is an IgG Fc polypeptide that differs from a reference IgG Fc polypeptide by single or multiple amino acid substitutions, deletions, and/or additions and substantially retains at least one biological activity of the reference IgG Fc polypeptide.
[0292] An "amino acid derivative," as used herein, refers to any amino acid, modified amino acid, and/or amino acid analogue, that is not one of the 20 common natural amino acids found in humans. Exemplary amino acid derivatives include natural amino acids not found in humans (e.g., seleno cysteine and pyrolysine, which may be found in some microorganisms) and unnatural amino acids. Exemplary amino acid derivatives, include, but are not limited to, amino acid derivatives commercially available through chemical product manufacturers (e.g., sigmaaldrich.com/chemistry/chemistry-products.html?TablePage=16274965, accessed on May 6, 2017, which is incorporated herein by reference). One or more amino acid derivatives may be incorporated into a polypeptide at a specific location using a translation system that utilizes host cells, orthogonal aminoacyl-tRNA synthetases derived from eubacterial synthetases, orthogonal tRNAs, and an amino acid derivative. For further descriptions, see, e.g., U.S. Pat. No. 9,624,485.
[0293] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution with an amino acid derivative. In some embodiments, the amino acid derivative is an alanine derivative, a cysteine derivative, an aspartic acid derivative, a glutamic acid derivative, a phenylalanine derivative, a glycine derivative, a histidine derivative, an isoleucine derivative, a lysine derivative, a leucine derivative, a methionine derivative, an asparagine derivative, a proline derivative, a glutamine derivative, an arginine derivative, a serine derivative, a threonine derivative, a valine derivative, a tryptophan derivative, or a tyrosine derivative.
[0294] In some embodiments, a variant IgG Fc polypeptide comprises a variant IgG Fc polypeptide of a companion animal species. In some embodiments, a variant IgG Fc polypeptide comprises a variant canine IgG Fc polypeptide, a variant equine IgG Fc polypeptide, or a feline IgG Fc polypeptide.
Exemplary Variant IgG Fc Polypeptides with Modified Protein a Binding
[0295] In some embodiments, a variant IgG Fc polypeptide has modified Protein A binding affinity. In some embodiments, a variant IgG Fc polypeptide has increased binding affinity to Protein A. In some embodiments, a variant IgG Fc polypeptide may be purified using Protein A column chromatography.
[0296] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 21, position 23, position 25, position 80, position 205, and/or position 207 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 21, position 23, and/or position 24 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 21, position 23, position 25, position 80, and/or position 207 of SEQ ID NO: 4.
[0297] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 15, and/or position 203 of SEQ ID NO: 64. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 199 and/or position 200 of SEQ ID NO: 67. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 199, position 200, position 201, and/or 202 of SEQ ID NO: 68.
[0298] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 21, position 23, position 25, position 80, position 205, and/or position 207 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 21, position 23, and/or position 24 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 21, position 23, position 25, position 80, and/or position 207 of SEQ ID NO: 3.
[0299] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 15 and/or position 203 of SEQ ID NO: 64. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 199 and/or position 200 of SEQ ID NO: 67. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 199, position 200, position 201, and/or position 202 of SEQ ID NO: 68.
[0300] In some embodiments, a variant IgG Fc polypeptide comprises a threonine at a position corresponding to position 21 of SEQ ID NO: 1, a leucine at a position corresponding to position 23 of SEQ ID NO: 1, an alanine at a position corresponding to position 25 of SEQ ID NO: 1, a glycine at a position corresponding to position 80 of SEQ ID NO: 1, an alanine at a position corresponding to position 205 of SEQ ID NO: 1, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises a threonine at a position corresponding to position 21 of SEQ ID NO: 3, a leucine at a position corresponding to position 23 of SEQ ID NO: 3, and/or an isoleucine at a position corresponding to position 24 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a threonine at a position corresponding to position 21 of SEQ ID NO: 4, a leucine at a position corresponding to position 23 of SEQ ID NO: 4, an alanine at a position corresponding to position 25 of SEQ ID NO: 4, a glycine at a position corresponding to position 80 of SEQ ID NO: 3, and/or a histidine at a position corresponding to position 207 of SEQ ID NO: 4.
[0301] In some embodiments, a variant IgG Fc polypeptide comprises a threonine or a valine at a position corresponding to position 15 of SEQ ID NO: 64, and/or a tyrosine or a valine at a position corresponding to position 203 of SEQ ID NO: 64. In some embodiments, a variant IgG Fc polypeptide comprises a leucine at a position corresponding to position 199 of SEQ ID NO: 67, and/or a histidine at a position corresponding to position 200 of SEQ ID NO: 67. In some embodiments, a variant IgG Fc polypeptide comprises an isoleucine at a position corresponding to position 199 of SEQ ID NO: 68, a histidine at a position corresponding to position 200 of SEQ ID NO: 68, an asparagine at a position corresponding to position 201 of SEQ ID NO: 68, and/or a histidine at a position corresponding to position 202 of SEQ ID NO: 68.
[0302] In some embodiments, a variant IgG Fc polypeptide comprises a threonine at position 21 of SEQ ID NO: 1, a leucine at position 23 of SEQ ID NO: 1, an alanine at position 25 of SEQ ID NO: 1, a glycine at position 80 of SEQ ID NO: 1, an alanine at position 205 of SEQ ID NO: 1, and/or a histidine at position 207 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises a threonine at position 21 of SEQ ID NO: 3, a leucine at position 23 of SEQ ID NO: 3, and/or an isoleucine at position 24 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprise a threonine at a position 21 of SEQ ID NO: 4, a leucine at position 23 of SEQ ID NO: 4, an alanine at position 25 of SEQ ID NO: 4, a glycine at position 80 of SEQ ID NO: 4, and/or a histidine at position 207 of SEQ ID NO: 4.
[0303] In some embodiments, a variant IgG Fc polypeptide comprises a threonine or a valine at position 15 of SEQ ID NO: 64, and/or a tyrosine or a valine at position 203 of SEQ ID NO: 64. In some embodiments, a variant IgG Fc polypeptide comprises a leucine at position 199 of SEQ ID NO: 67, and/or a histidine at position 200 of SEQ ID NO: 67. In some embodiments, a variant IgG Fc polypeptide comprises an isoleucine at position 199 of SEQ ID NO: 68, a histidine at position 200 of SEQ ID NO: 68, an asparagine at position 201 of SEQ ID NO: 68, and/or a histidine at position 202 of SEQ ID NO: 68.
[0304] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 60, SEQ ID NO: 61, SEQ ID NO: 62, or SEQ ID NO: 84. In some embodiments, a variant IgG Fc polypeptide comprises SEQ ID NO: 19, SEQ ID NO: 71, SEQ ID NO: 72, SEQ ID NO: 75, or SEQ ID NO: 76.
Exemplary Variant IgG Fc Polypeptides with Modified CD16 Binding
[0305] In some embodiments, a variant IgG Fc polypeptide has modified CD16 binding affinity. In some embodiments, a variant IgG Fc polypeptide has decreased binding affinity to CD16. In some embodiments, a variant IgG Fc may have a reduced ADCC immune response.
[0306] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 5, position 38, position 39, position 97, and/or position 98 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 5, position 38, position 39, position 97, and/or position 98 of SEQ ID NO: 3.
[0307] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 5, position 38, position 39, position 97, and/or position 98 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 5, position 38, position 39, position 97, and/or position 98 of SEQ ID NO: 3.
[0308] In some embodiments, a variant IgG Fc polypeptide comprises a proline at a position corresponding to position 5, a glycine at a position corresponding to position 38, an arginine at a position corresponding to position 39, a isoleucine at a position corresponding to position 97, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises a proline at a position corresponding to position 5, a glycine at a position corresponding to position 38, an arginine at a position corresponding to position 39, a isoleucine at a position corresponding to position 97, and/or a glycine at a position corresponding to position 98 of SEQ ID NO: 3.
[0309] In some embodiments, a variant IgG Fc polypeptide comprises a proline at position 5, a glycine at position 38, an arginine at position 39, a isoleucine at position 97, and/or a glycine at position 98 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises a proline at position 5, a glycine at position 38, an arginine at position 39, a isoleucine at position 97, and/or a glycine at position 98 of SEQ ID NO: 3.
[0310] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid sequence of SEQ ID NO: 139, SEQ ID NO: 140, SEQ ID N: 141, SEQ ID NO: 142, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 145, SEQ ID NO: 146, SEQ ID NO: 147, SEQ ID NO: 148, SEQ ID NO: 149, SEQ ID NO: 150, SEQ ID NO: 151, SEQ ID NO: 152, SEQ ID NO: 154, SEQ ID NO: 155, SEQ ID NO: 156, or SEQ ID NO: 157.
Exemplary Variant IgG Fc Polypeptides with Modified C1q Binding
[0311] In some embodiments, a variant IgG Fc polypeptide has modified C1q binding affinity. In some embodiments, a variant IgG Fc polypeptide has reduced binding affinity to C1q. In some embodiments, a variant IgG Fc polypeptide may have reduced complement fixation. In some embodiments, a variant IgG Fc may have a reduced complement-mediated immune response.
[0312] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 93 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 63. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 65. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 87 of SEQ ID NO: 66. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 198 of SEQ ID NO: 80. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 198 of SEQ ID NO: 81.
[0313] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 93 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 93 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 87 of SEQ ID NO: 63. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 87 of SEQ ID NO: 65. In some embodiments, a variant IgG Fc polypeptide comprises or an amino acid substitution at position 87 of SEQ ID NO: 66. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 198 of SEQ ID NO: 80. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 198 of SEQ ID NO: 81.
[0314] In some embodiments, a variant IgG Fc polypeptide comprises an arginine at a position corresponding to position 93 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises an arginine at a position corresponding to position 93 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 87 of SEQ ID NO: 63. In some embodiments, a variant IgG Fc polypeptide comprises a serine substitution at a position corresponding to position 87 of SEQ ID NO: 65. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 87 of SEQ ID NO: 66. In some embodiments, a variant IgG Fc polypeptide comprises an alanine at a position corresponding to position 198 of SEQ ID NO: 80. In some embodiments, a variant IgG Fc polypeptide comprises an alanine at a position corresponding to position 198 of SEQ ID NO: 81.
[0315] In some embodiments, a variant IgG Fc polypeptide comprises an arginine at position 93 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 93 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 87 of SEQ ID NO: 63. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 87 of SEQ ID NO: 65. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 87 of SEQ ID NO: 66. In some embodiments, a variant IgG Fc polypeptide comprises an alanine at position 198 of SEQ ID NO: 80. In some embodiments, a variant IgG Fc polypeptide comprises an alanine at position 198 of SEQ ID NO: 81.
[0316] In some embodiments, a variant IgG Fc polypeptide comprises the amino acid sequence of SEQ ID NO: 78, SEQ ID NO: 79, or SEQ ID NO: 84. In some embodiments, a variant IgG Fc polypeptide comprises the amino acid sequence of SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 74, or SEQ ID NO: 77. In some embodiments, a variant IgG Fc polypeptide comprises the amino acid sequence of SEQ ID NO: 82 or SEQ ID NO: 83.
Exemplary Variant IgG Fc Polypeptides with a Modified Inter-Chain Disulfide Linkage
[0317] In some embodiments, a variant feline IgG Fc polypeptide has at least one additional inter-chain disulfide linkage relative to the wild-type feline IgG Fc polypeptide. In some embodiments, a variant feline IgG Fc polypeptide has at least one additional inter-chain disulfide linkage in the hinge region. In some embodiments, a variant feline IgG2 Fc polypeptide with at least one additional inter-chain disulfide linkage has increased inter-chain stability relative to the wild-type feline IgG Fc polypeptide. In some embodiments, a variant IgG polypeptide has at least one amino acid modification to a hinge region relative to a wild-type IgG Fc polypeptide. In some embodiments, the wild-type IgG Fc polypeptide is a wild-type feline or equine IgG Fc polypeptide. In some embodiments, the variant IgG Fc polypeptide comprises a hinge region or a portion of a hinge region from an IgG Fc polypeptide of a different isotype. In some embodiments, the variant IgG Fc polypeptide comprises a hinge region from a wild-type feline IgG-1a Fc polypeptide, from a wild-type feline IgG-1b Fc polypeptide, or from a wild-type equine IgG1 Fc polypeptide. In some embodiments, a variant IgG2 Fc polypeptide has increased recombinant production and/or increased hinge disulfide formation relative to the wild-type IgG Fc polypeptide. In some embodiments, the increased recombinant production and/or increased hinge disulfide formation can be determined by SDS-PAGE analysis under reducing and/or non-reducing conditions.
[0318] In some embodiments, a variant IgG Fc polypeptide comprises a cysteine at a position corresponding to position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, or position 16 of SEQ ID NO: 16. In some embodiments, a variant IgG Fc polypeptide comprises a cysteine at position 8, position 9, position 10, position 11, position 12, position 13, position 14, position 15, or position 16 of SEQ ID NO: 16. In some embodiments, a variant IgG Fc polypeptide comprises SEQ ID NO: 17.
[0319] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at a position corresponding to position 3 and/or at a position corresponding to position 20 of SEQ ID NO: 129.
[0320] In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118. In some embodiments, a variant IgG Fc polypeptide comprises an amino acid substitution at position 3 and/or at a position corresponding to position 20 of SEQ ID NO: 129.
[0321] In some embodiments, a variant IgG Fc polypeptide comprises a proline at a position corresponding to position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 3 and/or a proline at a position corresponding to position 20 of SEQ ID NO: 129.
[0322] In some embodiments, a variant IgG Fc polypeptide comprises a proline at position 16 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 3 and/or a proline at position 20 of SEQ ID NO: 129.
[0323] In some embodiments, the variant IgG Fc polypeptide comprises SEQ ID NO: 19, SEQ ID NO: 125 or SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 128, SEQ ID NO: SEQ ID NO: 129, SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 133, SEQ ID N: 134, SEQ ID NO: 135.
Exemplary Variant IgG Fc Polypeptides for Heterodimeric Polypeptides
[0324] In certain embodiments, a heterodimeric polypeptide provided herein is a a bispecific antibody. A bispecific antibody has a binding specificity for two different epitopes or target molecules. In some embodiments, a bispecific antibody binds to two different epitopes of the same target molecule. Bispecific antibodies may be full length antibodies or antibody fragments.
[0325] In some embodiments, the heterodimeric polypeptide comprises a first variant IgG Fc polypeptide comprising a "knob" mutation and a second variant IgG Fc polypeptide comprising a "hole" mutation. Nonlimiting exemplary knob and hole mutations are described, for example, in Merchant, A. M. et al. An efficient route to human bispecific IgG. Nat Biotechnol, 16(7):677-81 (1998).
[0326] In some embodiments, a variant canine or variant feline IgG Fc polypeptide comprises a knob mutation. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at a position corresponding to position 138 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at a position corresponding to position 137 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at a position corresponding to position 137 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at a position corresponding to position 138 of SEQ ID NO: 4. In some embodiments, a variant IgG Fc polypeptide comprises a tryptophan at a position corresponding to position 154 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118.
[0327] In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at position 138 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at position 137 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at position 137 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a tyrosine or a tryptophan at position 138 of SEQ ID NO: 4. In some embodiments, a variant IgG Fc polypeptide comprises a tryptophan at position 154 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118.
[0328] In some embodiments, a variant IgG Fc polypeptide comprising a knob mutation comprises an amino acid sequence of SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, SEQ ID NO: 14, SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, SEQ ID NO: 115, SEQ ID NO: 119, SEQ ID NO: 121, or SEQ ID NO: 123.
[0329] In some embodiments, a variant canine or a variant feline IgG Fc polypeptide comprises a hole mutation. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 138, an alanine at a position corresponding to position 140, and/or a threonine at a position corresponding to position 181 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 137, an alanine at a position corresponding to position 139, and/or a threonine at a position corresponding to position 180 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 137, an alanine at a position corresponding to position 139, and/or a threonine at a position corresponding to position 180 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 138, an alanine at a position corresponding to position 140, and/or a threonine at a position corresponding to position 181 of SEQ ID NO: 4. In some embodiments, a variant IgG Fc polypeptide comprises a serine at a position corresponding to position 154, an alanine at a position corresponding to position 156, and/or a threonine at a position corresponding to position 197 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118.
[0330] In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 138, an alanine at position 140, and/or a threonine at position 181 of SEQ ID NO: 1. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 137, an alanine at position 139, and/or a threonine at position 181 of SEQ ID NO: 2. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 137, an alanine at position 139, and/or a threonine at position 181 of SEQ ID NO: 3. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 138, an alanine at position 140, and/or a threonine at position 181 of SEQ ID NO: 4. In some embodiments, a variant IgG Fc polypeptide comprises a serine at position 154, an alanine at position 156, and/or a threonine at position 197 of SEQ ID NO: 16, SEQ ID NO: 80, SEQ ID NO: 81, SEQ ID NO: 117, or SEQ ID NO: 118.
[0331] In some embodiments, a variant IgG Fc polypeptide comprising a hole mutation comprises an amino acid sequence of SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, SEQ ID NO: 15, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 116, SEQ ID NO: 120, SEQ ID NO: 122, or SEQ ID NO: 124.
[0332] In some embodiments, a contiguous polypeptide comprises a GLP1 polypeptide and a variant canine or feline IgG Fc polypeptide comprising a knob mutation. In some embodiments, a contiguous polypeptide comprises a glucagon polypeptide and a variant canine or feline IgG Fc polypeptide comprising a knob mutation. In some embodiments, a contiguous polypeptide comprises a GLP1 polypeptide and a variant canine or feline IgG Fc polypeptide comprising a hole mutation. In some embodiments, a contiguous polypeptide comprises a glucagon polypeptide and a variant canine or feline IgG Fc polypeptide comprising a hole mutation.
[0333] In some embodiments, the heterodimeric polypeptide comprises a first contiguous polypeptide comprises a GLP1 polypeptide and a variant canine or feline IgG Fc polypeptide comprising a knob mutation, and a second contiguous polypeptide comprises a glucagon polypeptide and a variant canine or feline IgG Fc polypeptide comprising a hole mutation. In some embodiments, the heterodimeric polypeptide comprises a first contiguous polypeptide comprises a glucagon polypeptide and a variant canine or feline IgG Fc polypeptide comprising a knob mutation, and a second contiguous polypeptide comprises a GLP1 polypeptide and a variant canine or feline IgG Fc polypeptide comprising a hole mutation.
Exemplary GLP1 Polypeptides and Glucagon Polypeptides
[0334] "GLP1" or a "GLP1 polypeptide," as used herein, is a polypeptide comprising the entirety or a fragment of glucagon-like peptide-1 that binds to a glucagon-like peptide 1 receptor (GLP1R).
[0335] For example, "GLP1" refers to a GLP1 polypeptide from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys), rodents (e.g., mice and rats), and companion animals (e.g., dogs, cats, and equine), unless otherwise indicated. In some embodiments, a GLP1 polypeptide is a wild-type GLP1 polypeptide, such as GLP1 (7-37) (SEQ ID N: 85). In some embodiments, a GLP1 polypeptide is a variant GLP1 polypeptide, such as GLP1 (7-36) (SEQ ID NO: 98), GLP1 (7-35) (SEQ ID NO: 99), GLP1-S8 (7-35) (SEQ ID NO: 86), or GLP1-G8 (7-35) (SEQ ID NO: 87). In some embodiments, GLP1 comprises the amino acid sequence of SEQ ID NO: 20.
[0336] "GLP1R," as used herein, is a polypeptide comprising the entirety or a fragment of a glucagon-like peptide 1 receptor that is capable of binding to a wild-type GLP1.
[0337] For example, "GLP1R" refers to a GLP1R polypeptide from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys), rodents (e.g., mice and rats), and companion animals (e.g., dogs, cats, and equine), unless otherwise indicated. In some embodiments, GLP1R is an extracellular domain fragment that binds a wild-type GLP1 polypeptide. In some embodiments, a feline GLP1R comprises the amino acid sequence of SEQ ID NO: 49. In some embodiments, a feline GLP1R comprises the amino acid sequence of SEQ ID NO: 48.
[0338] An "extracellular domain" ("ECD") is the portion of a polypeptide that extends beyond the transmembrane domain into the extracellular space. The term "extracellular domain," as used herein, may comprise a complete extracellular domain or may comprise a truncated extracellular domain missing one or more amino acids, that binds to its ligand. The composition of the extracellular domain may depend on the algorithm used to determine which amino acids are in the membrane. Different algorithms may predict, and different systems may express, different extracellular domains for a given protein.
[0339] "Glucagon" or a "glucagon polypeptide," as used herein, is a polypeptide comprising the entirety or a fragment of glucagon that binds to a glucagon receptor.
[0340] For example, "glucagon" refers to a glucagon polypeptide from any vertebrate source, including mammals such as primates (e.g., humans and cynomolgus monkeys), rodents (e.g., mice and rats), and companion animals (e.g., dogs, cats, and equine), unless otherwise indicated. In some embodiments, a glucagon polypeptide is a wild-type glucagon polypeptide, such as SEQ ID NO: 21.
Exemplary Variant IgG Fc Polypeptides and Fusion Molecules
[0341] Polypeptides and other molecules may comprise a variant IgG Fc polypeptide. In some embodiments, a fusion molecule comprises a variant IgG Fc polypeptide, such as the variant IgG Fc polypeptides described herein. In some embodiments, an antibody or an antibody fragment comprises a variant IgG Fc polypeptide, such as the variant IgG Fc polypeptides described herein.
[0342] A "fusion molecule," as used herein, refers to a molecule comprising one or more "fusion partners." In some embodiments, the fusion partners are covalently linked ("fused"). If two fusion partners are both polypeptides, the fusion partner polypeptides may be part of a contiguous amino acid sequence (i.e., a contiguous polypeptide). A first fusion partner polypeptide may be linked to either the N-terminus or the C-terminus of a second fusion partner. In some embodiments, the fusion partners are translated as a single polypeptide from a coding sequence that encodes both fusion partners. Fusion partners may be covalently linked through other means, such as, for example, a chemical linkage other than a peptide bond. Many known methods of covalently linking polypeptides to other molecules (for example, fusion partners) may be used. In other embodiments, the fusion partners are fused through a "linker," which is comprised of at least one amino acid or chemical moiety. In some embodiments, fusion partners are noncovalently linked. In some such embodiments, they may be linked, for example, using binding pairs. Exemplary binding pairs include, but are not limited to, biotin and avidin or streptavidin, an antibody and its antigen, etc.
[0343] In some embodiments, the fusion partners include an IgG Fc polypeptide and at least one GLP1 polypeptide. In some embodiments, the fusion partners include an IgG Fc polypeptide, a GLP1 polypeptide, and a glucagon polypeptide. In some embodiments, a GLP1 polypeptide may be linked to either the N-terminus or the C-terminus of an IgG Fc polypeptide. In some embodiments, a glucagon polypeptide may be linked to either the N-terminus or the C terminus of an IgG Fc polypeptide.
[0344] The term "contiguous polypeptide" herein is used to mean an uninterrupted sequence of amino acids. A contiguous polypeptide is typically translated from a single continuous DNA sequence. It can be made by genetic engineering, for example, by removing the stop codon from the DNA sequence of the first protein, then appending the DNA sequence of the second protein in frame, so that the DNA sequence is expressed as a single protein. Typically, this is accomplished by cloning a cDNA into an expression vector in frame with an existing gene.
[0345] A "linker" refers to one or more amino acid residues that connects a first polypeptide with a second polypeptide.
[0346] In some embodiments, the linker is a flexible, non-structural linker. In some embodiments, the linker is a glycine-rich, serine-rich, or glycine- and serine-rich linker. In some embodiments, a linker comprises 100%, at least 95%, at least 90%, or at least 85% serine and/or glycine amino acid residues.
[0347] An "extension," as used herein, refers to one or more amino acid residues that are connected to a polypeptide at its C-terminus or at its N-terminus.
[0348] In some embodiments, an extension is flexible. In some embodiments, the extension adds flexibility to the polypeptide without interfering with the biological activity of the polypeptide. In some embodiments, the extension increases solubility of the polypeptide. In some embodiments, the extension comprises one or more glycine residues. In some embodiments, the extension comprises a glycine residue (SEQ ID NO: 88), two glycine residues (SEQ ID NO: 89), a three glycine residues (SEQ ID NO: 90), four glycine residues (SEQ ID NO: 91), five glycine residues (SEQ ID NO: 92), six glycine residues (SEQ ID NO: 93), seven glycine residues (SEQ ID NO: 94), eight glycine residues (SEQ ID NO: 95), or more glycine residues.
[0349] In some embodiments, the contiguous polypeptide comprises an IgG Fc polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 100, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 167, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, or 199 and a GLP1 polypeptide comprising an amino acid sequence of SEQ ID NO: 85. In some embodiments, the contiguous polypeptide comprises an IgG Fc polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 100, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 167, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, or 199 and a GLP1 polypeptide comprising an amino acid sequence of SEQ ID NO: 86. In some embodiments, the contiguous polypeptide comprises an IgG Fc polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 100, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 167, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, or 199 and a GLP1 polypeptide comprising an amino acid sequence of SEQ ID NO: 87. In some embodiments, the contiguous polypeptide comprises an IgG Fc polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 100, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 167, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, or 199 and a GLP1 polypeptide comprising an amino acid sequence of SEQ ID NO: 98. In some embodiments, the contiguous polypeptide comprises an IgG Fc polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 100, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 167, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, or 199 and a GLP1 polypeptide comprising an amino acid sequence of SEQ ID NO: 99.
[0350] In some embodiments, the contiguous polypeptide comprises an IgG Fc polypeptide comprising an amino acid sequence of any one of SEQ ID NOs: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 100, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 167, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, or 199 and a glucagon polypeptide comprising an amino acid sequence of SEQ ID NO: 21.
[0351] In some embodiments, a contiguous polypeptide comprises a first GLP1 polypeptide (GLP1A), a first linker (L1), an Fc polypeptide of a companion animal species, optionally a second linker (L2), and optionally a second GLP1 polypeptide (GLP1B). In some embodiments, the contiguous polypeptide comprises:
GLP1A-L1-Fc; Formula (I):
Fc-L1-GLP1A; or Formula (II):
GLP1A-L1-Fc-L2-GLP1B. Formula (III):
[0352] In some embodiments, a contiguous polypeptide comprises a GLP1 polypeptide, a first linker (L1), an Fc polypeptide, a second linker (L2), and a glucagon polypeptide (Gluc). In some embodiments, the contiguous polypeptide comprises:
GLP1-L1-Fc-L2-Gluc; or Formula (IV):
Gluc-L1-Fc-L2-GLP1. Formula (V):
[0353] In some embodiments, the GLP1 fusion molecule has an increased serum half-life compared to a wild-type GLP1 polypeptide. The increased half-life of the GLP1 fusion molecules described herein may require lower doses and less-frequent dosing regimen than wild-type GLP1 polypeptides.
[0354] In some embodiments, GLP1B, if present, comprises the same amino acid sequence as GLP1A.
[0355] In some embodiments, GLP1, GLP1A, or GLP1B, if present, comprises a wild-type GLP1 polypeptide. In some embodiments, GLP1, GLP1A, or GLP1B, if present, comprises a variant GLP1 polypeptide. In some embodiments, GLP1, GLP1A, or GLP1B, if present, comprises an amino acid sequence of SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 98, or SEQ ID NO: 99.
[0356] In some embodiments, the glucagon polypeptide comprises a wild-type glucagon polypeptide. In some embodiments, the glucagon polypeptide comprises an amino acid sequence of SEQ ID NO: 21. In some embodiments, the glucagon polypeptide comprises a variant glucagon polypeptide.
[0357] In some embodiments, the Fc polypeptide is a human IgG Fc. In some embodiments, the Fc polypeptide is a human IgG1 Fc, a human IgG2 Fc, a human IgG3 Fc, or a human IgG4 Fc. In some embodiments, the Fc polypeptide is a variant human IgG Fc.
[0358] In some embodiments, the Fc polypeptide is an IgG Fc from a companion animal.
[0359] In some embodiments, the Fc polypeptide is a canine IgG-A Fc, a canine IgG-B Fc, a canine IgG-C Fc, a canine IgG-D Fc. In some embodiments, the Fc is an equine IgG1 Fc, an equine IgG2 Fc, an equine IgG3 Fc, an equine IgG4 Fc, an equine IgG5 Fc, an equine IgG6 Fc, or an equine IgG7 Fc. In some embodiments, the Fc is a feline IgG1a Fc, a feline IgG1b Fc, or a feline IgG2 Fc.
[0360] In some embodiments, the Fc polypeptide is a variant IgG Fc. In some embodiments, the FC polypeptide is a variant canine IgG-A Fc, a variant canine IgG-B Fc, a variant canine IgG-C Fc, a variant canine IgG-D Fc. In some embodiments, the Fc is a variant equine IgG1 Fc, a variant equine IgG2 Fc, a variant equine IgG3 Fc, a variant equine IgG4 Fc, a variant equine IgG5 Fc, a variant equine IgG6 Fc, or a variant equine IgG7 Fc. In some embodiments, the Fc is a variant feline IgG1a Fc, a variant feline IgG1b Fc, or a variant feline IgG2 Fc.
[0361] In some embodiments, L1 and L2, if present, each independently is a flexible linker. In some embodiments, the amino acid sequence of L1 and L2, if present, each independently comprises 100%, at least 95%, at least 90%, at least 85% serine and/or glycine amino acid residues.
[0362] In some embodiments, the contiguous polypeptide comprises an extension at its C-terminus. In some embodiments, the contiguous polypeptide comprises a glycine residue, two glycine residues, three glycine residues, four glycine residues, five glycine residues, six glycine residues, seven glycine residues, eight glycine residues, or greater than eight glycine residues at its C-terminus. In some embodiments, the contiguous polypeptide comprises an amino acid sequence of SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, or SEQ ID NO: 95 at its C-terminus.
[0363] In some embodiments, the contiguous polypeptide comprises the amino acid sequence of SEQ ID NO: 23; SEQ ID NO: 24; SEQ ID NO: 25; SEQ ID NO: 26; SEQ ID NO: 27; SEQ ID NO: 28; SEQ ID NO: 29; SEQ ID NO: 30; SEQ ID NO: 31; SEQ ID NO: 32; SEQ ID NO: 33; SEQ ID NO: 34; SEQ ID NO: 35; SEQ ID NO: 36; SEQ ID NO: 37; SEQ ID NO: 38; SEQ ID NO: 39; SEQ ID NO: 40; SEQ ID NO: 41; SEQ ID NO: 42; SEQ ID NO: 43; SEQ ID NO: 44; SEQ ID NO: 45; SEQ ID NO: 46; SEQ ID NO: 47, SEQ ID NO: 96, SEQ ID NO: 97, SEQ ID NO: 52; SEQ ID NO: 53; SEQ ID NO: 54; SEQ ID NO: 55; SEQ ID NO: 56; SEQ ID NO: 57; SEQ ID NO: 58; SEQ ID NO: 59; SEQ ID NO: 103; SEQ ID NO: 104; SEQ ID NO: 105; or SEQ ID NO: 106.
[0364] A nucleotide sequence encoding a polypeptide of interest, such as a variant IgG Fc polypeptide or other polypeptide described herein, can be inserted into an expression vector suitable for expression in a selected host cell. A variant IgG Fc polypeptide or other polypeptide described herein may be expressed by culturing a host cell transfected with an expression vector comprising the nucleotide sequence.
[0365] A "vector" is a plasmid that can be used to transfer DNA sequences from one organism to another or to express a gene of interest. A vector typically includes an origin of replication and regulatory sequences which regulate the expression of the gene of interest, and may or may not carry a selective marker gene, such as an antibiotic resistance gene. A vector is suitable for the host cell in which it is to be expressed. A vector may be termed a "recombinant vector" when the gene of interest is present in the vector.
[0366] A "host cell" refers to a cell that may be or has been a recipient of a vector or isolated polynucleotide. Host cells may be prokaryotic cells or eukaryotic cells. Exemplary eukaryotic cells include mammalian cells, such as primate or non-primate animal cells; fungal cells, such as yeast; plant cells; and insect cells. Nonlimiting exemplary mammalian cells include, but are not limited to, NS0 cells, PER.C6.RTM. cells (Crucell), 293 cells, and CHO cells, and their derivatives, such as 293-6E, DG44, CHO-S, and CHO-K cells. Host cells include progeny of a single host cell, and the progeny may not necessarily be completely identical (in morphology or in genomic DNA complement) to the original parent cell due to natural, accidental, or deliberate mutation. A host cell includes cells transfected in vivo with a polynucleotide(s) encoding an amino acid sequence(s) provided herein.
[0367] The term "isolated" as used herein refers to a molecule that has been separated from at least some of the components with which it is typically found in nature or produced. For example, a polypeptide is referred to as "isolated" when it is separated from at least some of the components of the cell in which it was produced. Where a polypeptide is secreted by a cell after expression, physically separating the supernatant containing the polypeptide from the cell that produced it is considered to be "isolating" the polypeptide. Similarly, a polynucleotide is referred to as "isolated" when it is not part of the larger polynucleotide (such as, for example, genomic DNA or mitochondrial DNA, in the case of a DNA polynucleotide) in which it is typically found in nature, or is separated from at least some of the components of the cell in which it was produced, for example, in the case of an RNA polynucleotide. Thus, a DNA polynucleotide that is contained in a vector inside a host cell may be referred to as "isolated."
[0368] A "signal sequence" refers to a sequence of amino acid residues or polynucleotides encoding such, which facilitates secretion of a polypeptide of interest and is typically cleaved upon export of the polypeptide to the outside of the cell surface membrane.
[0369] In some embodiments, a variant IgG Fc polypeptide or a contiguous polypeptide comprising a variant Fc polypeptide is isolated using chromatography, such as size exclusion chromatography, ion exchange chromatography, protein A column chromatography, hydrophobic interaction chromatography, and CHT chromatography.
[0370] A label can be attached to a variant IgG Fc polypeptides or a contiguous polypeptide comprising a variant Fc polypeptide. A "label" means a moiety attached to a molecule to render it detectable. In some embodiments, a variant IgG Fc polypeptide or a contiguous polypeptide comprising a variant Fc polypeptide is labeled with a detectable moiety including but not limited to radioisotopes, fluorescent labels, and various enzyme-substrate labels known in the art. In some embodiments, the label is a detectable marker that can produce a signal that is detectable by visual or instrumental means, for example, incorporation of a radiolabeled amino acid or attachment to a polypeptide of biotinyl moieties that can be detected by marked avidin (for example, streptavidin containing a fluorescent marker or enzymatic activity that can be detected by optical or colorimetric methods). Examples of labels for polypeptides include, but are not limited to, the following: radioisotopes or radionuclides (for example, .sup.3H, .sup.14C, .sup.35S, .sup.90Y, .sup.99Tc, .sup.11In, .sup.125I, .sup.131I, .sup.177Lu, .sup.166Ho, or .sup.153Sm); chromogens, fluorescent labels (for example, FITC, rhodamine, lanthanide phosphors), enzymatic labels (for example, p-galactosidase, horseradish peroxidase, luciferase, alkaline phosphatase); chemiluminescent markers; biotinyl groups; predetermined polypeptide epitopes recognized by a secondary reporter (for example, leucine zipper pair sequences, binding sites for secondary antibodies, metal binding domains, epitope tags); and magnetic agents, such as gadolinium chelates. Representative examples of labels commonly employed for immunoassays include moieties that produce light, for example, acridinium compounds, and moieties that produce fluorescence, for example, fluorescein. In this regard, the moiety itself may not be detectably labeled but may become detectable upon reaction with yet another moiety. General techniques to be used in performing the various immunoassays noted above are known to those of ordinary skill in the art.
Exemplary Variant IgG Fc Polypeptide Affinity to Protein A and/or C1q and/or CD16
[0371] The variant IgG Fc polypeptides described herein may have altered binding affinity to Protein A and/or C1q and/or CD16. In some embodiments, a variant IgG Fc polypeptide has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide. Such variant IgG Fc polypeptides may be purified by Protein A column chromatography. In some embodiments, a variant IgG Fc polypeptide has reduced binding affinity to C1q relative to the wild-type IgG Fc polypeptide. Such variant IgG Fc polypeptides may have reduced complement-mediated immune responses. In some embodiments, a variant IgG Fc polypeptide has reduced binding affinity to CD16 relative to the wild-type IgG Fc polypeptide. Such variant IgG Fc polypeptides may have reduced ADCC immune responses. In some embodiments, a variant IgG Fc polypeptide has increased binding affinity to Protein A relative to the wild-type IgG Fc polypeptide and/or has reduced binding affinity to C1q relative to the wild-type IgG Fc polypeptide and/or has reduced binding affinity to CD16 relative to the wild-type IgG Fc polypeptide.
[0372] "Protein A," as used herein, is a polypeptide comprising the entirety or a portion of Protein A that is capable of binding a wild-type canine IgG-B Fc, a wild-type equine IgG1 Fc, a wild-type equine IgG3 Fc, a wild-type equine IgG4 Fc, a wild-type equine IgG7 Fc, a wild-type feline IgG1a Fc, a wild-type feline IgG1b Fc, or a wild-type feline IgG2 Fc.
[0373] "C1q" or "C1q complex" is used interchangeably to refer to a protein complex involved in the complement system, or a portion thereof, that can bind a wild-type canine IgG-B Fc, a wild-type canine IgG-C Fc, a wild-type equine IgG1 Fc, a wild-type equine IgG3 Fc, a wild-type equine IgG4 Fc, a wild-type equine IgG7 Fc, a wild-type feline IgG1a Fc, or a wild-type feline IgG1b Fc.
[0374] "CD16," as used herein, is a polypeptide comprising the entirety or a portion of CD16 that is capable of binding a wild-type canine IgG-A Fc or a wild-type canine IgG-D Fc. The term "binds" to a substance is a term that is well understood in the art, and methods to determine such binding are also well known in the art. A molecule is said to exhibit "binding" if it reacts, associates with, or has affinity for a particular cell or substance and the reaction, association, or affinity is detectable by one or more methods known in the art, such as, for example, immunoblot, ELISA, KinEx A, biolayer interferometry (BLI), surface plasmon resonance devices, or etc.
[0375] "Protein A +," as used herein, means that the Fc polypeptide has Protein A binding affinity. In some embodiments, a Protein A+Fc polypeptide comprises at least one an amino acid modification that increases Protein A binding affinity.
[0376] "Protein A -," as used herein, means that the Fc polypeptide has low or no Protein A binding affinity.
[0377] "C1q+," as used herein, means that the Fc polypeptide has C1q binding affinity.
[0378] "C1q -," as used herein, means that the Fc polypeptide has low or no C1q binding affinity. In some embodiments, a C1q-Fc polypeptide has at least one an amino acid modification that reduces C1q binding affinity.
[0379] "CD16+," as used herein, means that the Fc polypeptide has CD16 binding affinity.
[0380] "CD16-," as used herein, means that the Fc polypeptide has low or no CD16 binding affinity. In some embodiments, a CD16- Fc polypeptide has at least one an amino acid modification that reduces CD16 binding affinity.
[0381] The term "affinity" means the strength of the sum total of noncovalent interactions between a single binding site of a molecule (for example, a receptor) and its binding partner (for example, a ligand). The affinity of a molecule X for its partner Y can generally be represented by the dissociation constant (Ku). Affinity can be measured by common methods known in the art, such as, for example, immunoblot, ELISA, KinEx A, biolayer interferometry (BLI), or surface plasmon resonance devices.
[0382] "Surface plasmon resonance" denotes an optical phenomenon that allows for the analysis of real-time biospecific interactions by detection of alterations in protein concentrations within a biosensor matrix, for example using the BIAcore.TM. system (BIAcore International AB, a GE Healthcare company, Uppsala, Sweden and Piscataway, N.J.). For further descriptions, see Jonsson et al. (1993) Ann. Biol. Clin. 51: 19-26.
[0383] "Biolayer interferometry" refers to an optical analytical technique that analyzes the interference pattern of light reflected from a layer of immobilized protein on a biosensor tip and an internal reference layer. Changes in the number of molecules bound to the biosensor tip cause shifts in the interference pattern that can be measured in real-time. A nonlimiting exemplary device for biolayer interferometry is an Octet.RTM. system (Pall ForteBio LLC). See, e.g., Abdiche et al., 2008, Anal. Biochem. 377: 209-277.
[0384] The terms "K.sub.D," "K.sub.d," "Kd" or "Kd value" as used interchangeably to refer to the equilibrium dissociation constant of a receptor-ligand interaction or antibody-antigen interaction.
[0385] In some embodiments, a variant IgG Fc polypeptide binds to Protein A with a dissociation constant (K.sub.D) of less than 5.times.10.sup.-6 M, less than 1.times.10.sup.-6 M, less than 5.times.10.sup.-7 M, less than 1.times.10.sup.-7 M, less than 5.times.10.sup.-8 M, less than 1.times.10.sup.-8 M, less than 5.times.10.sup.-9 M, less than 1.times.10.sup.-9 M, less than 5.times.10.sup.-10 M, less than 1.times.10.sup.-10 M, less than 5.times.10.sup.-11 M, less than 1.times.10.sup.-11 M, less than 5.times.10.sup.-12 M, or less than 1.times.10.sup.-12 M, as measured by biolayer interferometry.
[0386] In some embodiments, a variant IgG Fc polypeptide binds to C1q or CD16 with a dissociation constant (K.sub.D) of greater than 5.times.10.sup.-6 M, greater than 1.times.10.sup.-5 M, greater than 5.times.10.sup.-5 M, greater than 1.times.10.sup.-4 M, greater than 5.times.10.sup.-4 M, or greater than 1.times.10.sup.-3 M, as measured by biolayer interferometry.
[0387] In some embodiments, the K.sub.D of an IgG Fc polypeptide, such as a variant IgG Fc polypeptide, to Protein A or to C1q or to CD16 is measured by using biolayer interferometry assays using a biosensor, such as an Octet.RTM. System (Pall ForteBio LLC, Fremont, Calif.) according to the supplier's instructions. In brief, biotinylated Protein A or C1q or CD16 is bound to the sensor tip and the association of IgG Fc polypeptide is monitored for a specified time or until steady state is reached. Dissociation may be monitored for a specified time or until steady state is reached. A buffer only blank curve is subtracted to correct for any drift. The data are fit to a 2:1 binding model using ForteBio data analysis software to determine association rate constant (k.sub.on), dissociation rate constant (k.sub.off), and the K.sub.d. The equilibrium dissociation constant (K.sub.D) is calculated as the ratio of k.sub.off/k.sub.on. The term "k.sub.on" refers to the rate constant for association of a molecule X to its partner Y and the term "k.sub.off" refers to the rate constant for dissociation of a molecule X or partner Y from the molecule X/partner Y complex.
[0388] To "increase" or "stimulate" means to increase, improve, or augment an activity, function, or amount as compared to a reference. In some embodiments, by "increase" or "stimulate" is meant the ability to cause an overall increase of about 5% or greater, of about 10% or greater, of about 20% or greater, of about 30% or greater, of about 40% or greater, of about 50% or greater, of about 60% or greater, of about 70% or greater, of about 80% or greater, of about 90% or greater, of about 100% or greater, of about 125% or greater, of about 200% or greater relative to a reference value. In some embodiments, by "increase" or "stimulate" is meant the ability to cause an overall increase of about 5% to about 50%, of about 10% to about 20%, of about 50% to about 100%, of about 25% to about 70% relative to a reference value. In some embodiments, by "increase" or "stimulate" is meant the ability to cause an overall increase of 50% or greater. In some embodiments, by "increase" or "stimulate" is meant the ability to cause an overall increase of 75%, 85%, 90%, 95%, or greater. In some embodiments, the amount noted above is stimulated or increased over a period of time, relative to a control dose (such as a placebo) over the same period of time.
[0389] In some embodiments, a variant IgG Fc polypeptide is capable of binding to Protein A with an increased affinity of about 5% or greater, of about 10% or greater, of about 20% or greater, of about 30% or greater, of about 40% or greater, of about 50% or greater, of about 60% or greater, of about 70% or greater, of about 80% or greater, of about 90% or greater, of about 100% or greater, of about 125% or greater, of about 150% or greater, of about 200% or greater relative to a reference IgG Fc polypeptide. In some embodiments, a variant IgG Fc polypeptide is capable of binding to Protein A with an increased affinity of about 5% to about 50%, of about 10% to about 20%, of about 50% to about 100%, of about 25% to about 70% relative to a reference IgG Fc polypeptide. In some embodiments, the reference IgG Fc polypeptide is a wild-type IgG Fc polypeptide. In some embodiments, the reference IgG Fc polypeptide is a different variant IgG Fc polypeptide.
[0390] To "reduce" or "inhibit" means to decrease, reduce, or arrest an activity, function, or amount as compared to a reference. In some embodiments, by "reduce" or "inhibit" is meant the ability to cause an overall decrease of about 5% or greater, of about 10% or greater, of about 20% or greater, of about 30% or greater, of about 40% or greater, of about 50% or greater, of about 60% or greater, of about 70% or greater, of about 80% or greater, or of about 90% or greater relative to a reference IgG Fc polypeptide. In some embodiments, by "reduce" or "inhibit" is meant the ability to cause an overall decrease of about 5% to about 50%, of about 10% to about 20%, of about 50% to about 100%, of about 25% to about 70% relative to a reference value. In some embodiments, by "reduce" or "inhibit" is meant the ability to cause an overall decrease of 50% or greater. In some embodiments, by "reduce" or "inhibit" is meant the ability to cause an overall decrease of 75%, 85%, 90%, 95%, or greater. In some embodiments, the amount noted above is inhibited or decreased over a period of time, relative to a control dose (such as a placebo) over the same period of time.
[0391] In some embodiments, a variant IgG Fc polypeptide is capable of binding to C1q or CD16 with a decreased affinity of about 5% or greater, of about 10% or greater, of about 20% or greater, of about 30% or greater, of about 40% or greater, of about 50% or greater, of about 60% or greater, of about 70% or greater, of about 80% or greater, of about 90% or greater relative to a reference IgG Fc polypeptide. In some embodiments, a variant IgG Fc polypeptide is capable of binding to C1q or CD16 with a decreased affinity of about 5% to about 50%, of about 10% to about 20%, of about 50% to about 100%, of about 25% to about 70% relative to a reference IgG Fc polypeptide. In some embodiments, the reference IgG Fc polypeptide is a wild-type IgG Fc polypeptide. In some embodiments, the reference IgG Fc polypeptide is a different variant IgG Fc polypeptide.
[0392] A "reference" as used herein, refers to any sample, standard, or level that is used for comparison purposes. A reference may be a wild-type reference or a variant reference. A reference may be obtained from a healthy or non-diseased sample. In some examples, a reference is obtained from a non-diseased or non-treated sample of a companion animal. In some examples, a reference is obtained from one or more healthy animals of a particular species, which are not the animal being tested or treated.
Exemplary Biological Activity of Variant GLP1 Fusion Molecules
[0393] In some embodiments, a GLP1 fusion molecule binds to GLP1R and activates cAMP production. In some embodiments, a GLP1 fusion polypeptide increases production of cAMP in a cell by at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or 100% compared to signaling function in the absence of the GLP1 fusion polypeptide.
Exemplary Pharmaceutical Compositions
[0394] The terms "pharmaceutical formulation" and "pharmaceutical composition" refer to a preparation which is in such form as to permit the biological activity of the active ingredient(s) to be effective, and which contains no additional components that are unacceptably toxic to a subject to which the formulation would be administered.
[0395] A "pharmaceutically acceptable carrier" refers to a non-toxic solid, semisolid, or liquid filler, diluent, encapsulating material, formulation auxiliary, or carrier conventional in the art for use with a therapeutic agent that together comprise a "pharmaceutical composition" for administration to a subject. A pharmaceutically acceptable carrier is non-toxic to recipients at the dosages and concentrations employed and is compatible with other ingredients of the formulation. The pharmaceutically acceptable carrier is appropriate for the formulation employed. Examples of pharmaceutically acceptable carriers include alumina; aluminum stearate; lecithin; serum proteins, such as human serum albumin, canine or other animal albumin; buffers such as phosphate, citrate, tromethamine or HEPES buffers; glycine; sorbic acid; potassium sorbate; partial glyceride mixtures of saturated vegetable fatty acids; water; salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, or magnesium trisilicate; polyvinyl pyrrolidone, cellulose-based substances; polyethylene glycol; sucrose; mannitol; or amino acids including, but not limited to, arginine.
[0396] The pharmaceutical composition can be stored in lyophilized form. Thus, in some embodiments, the preparation process includes a lyophilization step. The lyophilized composition may then be reformulated, typically as an aqueous composition suitable for parenteral administration, prior to administration to the dog, cat, or horse. In other embodiments, particularly where a variant IgG Fc polypeptide or other polypeptide described herein is highly stable to thermal and oxidative denaturation, the pharmaceutical composition can be stored as a liquid, i.e., as an aqueous composition, which may be administered directly, or with appropriate dilution, to the dog, cat, or horse. A lyophilized composition can be reconstituted with sterile Water for Injection (WFI). Bacteriostatic reagents, such benzyl alcohol, may be included. Thus, the invention provides pharmaceutical compositions in solid or liquid form.
[0397] The pH of the pharmaceutical compositions may be in the range of from about pH 5 to about pH 8, when administered. The compositions of the invention are sterile if they are to be used for therapeutic purposes. Sterility can be achieved by any of several means known in the art, including by filtration through sterile filtration membranes (e.g., 0.2 micron membranes). Sterility may be maintained with or without anti-bacterial agents.
Certain Uses of Fc Polypeptides and Pharmaceutical Compositions
[0398] A polypeptide comprising a variant Fc polypeptide, such as a variant IgG Fc polypeptide, of the invention or pharmaceutical compositions comprising a variant Fc polypeptide of the invention may be useful for extending product half-life in vivo in a companion animal, including, but not limited to, canine, feline, or equine.
[0399] As used herein, "treatment" is an approach for obtaining beneficial or desired clinical results. "Treatment" as used herein, covers any administration or application of a therapeutic for disease in a mammal, including a companion animal. For purposes of this disclosure, beneficial or desired clinical results include, but are not limited to, any one or more of: alleviation of one or more symptoms, diminishment of extent of disease, preventing or delaying spread of disease, preventing or delaying recurrence of disease, delay or slowing of disease progression, amelioration of the disease state, inhibiting the disease or progression of the disease, inhibiting or slowing the disease or its progression, arresting its development, and remission (whether partial or total). Also encompassed by "treatment" is a reduction of pathological consequence of a proliferative disease. The methods provided herein contemplate any one or more of these aspects of treatment. In-line with the above, the term treatment does not require one-hundred percent removal of all aspects of the disorder.
[0400] A "therapeutically effective amount" of a substance/molecule, agonist or antagonist may vary according to factors such as the type of disease to be treated, the disease state, the severity and course of the disease, the type of therapeutic purpose, any previous therapy, the clinical history, the response to prior treatment, the discretion of the attending veterinarian, age, sex, and weight of the animal, and the ability of the substance/molecule, agonist or antagonist to elicit a desired response in the animal. A therapeutically effective amount is also one in which any toxic or detrimental effects of the substance/molecule, agonist or antagonist are outweighed by the therapeutically beneficial effects. A therapeutically effective amount may be delivered in one or more administrations. A therapeutically effective amount refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic or prophylactic result.
[0401] In some embodiments, a variant IgG Fc polypeptide or other polypeptide described herein, or a pharmaceutical composition comprising such is administered parenterally, by subcutaneous administration, intravenous infusion, or intramuscular injection. In some embodiments, a variant IgG Fc polypeptide or other polypeptide described herein, or a pharmaceutical composition comprising such is administered as a bolus injection or by continuous infusion over a period of time. In some embodiments, a variant IgG Fc polypeptide or other polypeptide described herein, or a pharmaceutical composition comprising such is administered by an intramuscular, an intraperitoneal, an intracerebrospinal, a subcutaneous, an intra-arterial, an intrasynovial, an intrathecal, or an inhalation route.
[0402] In some embodiments, a GLP1 fusion polypeptide or pharmaceutical compositions comprising it can be utilized in accordance with the methods herein to treat high blood glucose-related conditions. In some embodiments, an GLP1 fusion polypeptide or pharmaceutical compositions is administered to a companion animal, such as a canine, a feline, or equine, to treat high blood glucose-related condition.
[0403] In some embodiments, a variant IgG Fc polypeptide or other polypeptide described herein, or a pharmaceutical composition comprising such is administered in an amount in the range of 0.0001 mg/kg body weight to 100 mg/kg body weight per dose. In some embodiments, GLP1 analog may be administered in an amount in the range of 0.005 mg/kg body weight to 20 mg/kg body weight per dose. In some embodiments, GLP1 analog may be administered in an amount in the range of 1 mg/kg body weight to 10 mg/kg body weight per dose. In some embodiments, GLP1 analog may be administered in an amount in the range of 0.5 mg/kg body weight to 100 mg/kg body, in the range of 1 mg/kg body weight to 100 mg/kg body weight, in the range of 5 mg/kg body weight to 100 mg/kg body weight, in the range of 10 mg/kg body weight to 100 mg/kg body weight, in the range of 20 mg/kg body weight to 100 mg/kg body weight, in the range of 50 mg/kg body weight to 100 mg/kg body weight, in the range of 1 mg/kg body weight to 10 mg/kg body weight, in the range of 5 mg/kg body weight to 10 mg/kg body weight, in the range of 0.5 mg/kg body weight to 10 mg/kg body weight, or in the range of 5 mg/kg body weight to 50 mg/kg body weight.
[0404] In some embodiments, a variant IgG Fc polypeptide or other polypeptide described herein, or a pharmaceutical composition comprising such is administered to a companion animal at one time or over a series of treatments. In some embodiments, the dose is administered once per week for at least two or three consecutive weeks, and in some embodiments, this cycle of treatment is repeated two or more times, optionally interspersed with one or more weeks of no treatment. In other embodiments, the therapeutically effective dose is administered once per day for two to five consecutive days, and in some embodiments, this cycle of treatment is repeated two or more times, optionally interspersed with one or more days or weeks of no treatment.
[0405] Administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive or sequential administration in any order. The term "concurrently" is used herein to refer to administration of two or more therapeutic agents, where at least part of the administration overlaps in time or where the administration of one therapeutic agent falls within a short period of time relative to administration of the other therapeutic agent. For example, the two or more therapeutic agents are administered with a time separation of no more than about a specified number of minutes. The term "sequentially" is used herein to refer to administration of two or more therapeutic agents where the administration of one or more agent(s) continues after discontinuing the administration of one or more other agent(s), or wherein administration of one or more agent(s) begins before the administration of one or more other agent(s). For example, administration of the two or more therapeutic agents are administered with a time separation of more than about a specified number of minutes. As used herein, "in conjunction with" refers to administration of one treatment modality in addition to another treatment modality. As such, "in conjunction with" refers to administration of one treatment modality before, during or after administration of the other treatment modality to the animal.
[0406] In some embodiments, the dose is administered once per week for at least two or three consecutive weeks, and in some embodiments, this cycle of treatment is repeated two or more times, optionally interspersed with one or more weeks of no treatment. In other embodiments, the therapeutically effective dose is administered once per day for two to five consecutive days, and in some embodiments, this cycle of treatment is repeated two or more times, optionally interspersed with one or more days or weeks of no treatment.
[0407] Administration "in combination with" one or more further therapeutic agents includes simultaneous (concurrent) and consecutive or sequential administration in any order. The term "concurrently" is used herein to refer to administration of two or more therapeutic agents, where at least part of the administration overlaps in time or where the administration of one therapeutic agent falls within a short period of time relative to administration of the other therapeutic agent. For example, the two or more therapeutic agents are administered with a time separation of no more than about a specified number of minutes. The term "sequentially" is used herein to refer to administration of two or more therapeutic agents where the administration of one or more agent(s) continues after discontinuing the administration of one or more other agent(s), or wherein administration of one or more agent(s) begins before the administration of one or more other agent(s). For example, administration of the two or more therapeutic agents are administered with a time separation of more than about a specified number of minutes. As used herein, "in conjunction with" refers to administration of one treatment modality in addition to another treatment modality. As such, "in conjunction with" refers to administration of one treatment modality before, during or after administration of the other treatment modality to the animal.
[0408] In some embodiments, the method comprises administering in combination with a GLP1 fusion polypeptide insulin, a DPP4 inhibitor, a SGLT2 inhibitor, a biguanides sulfonylurea, a meglitinide derivative, an alpha-glucosidase inhibitor, a thiazolidinedione (TZD), an amylinomimetic, a bile acid sequestrant, a dopamine agonist.
[0409] The following examples illustrate particular aspects of the disclosure and are not intended in any way to limit the disclosure.
EXAMPLES
Example 1
Variant Canine IgG Fc Polypeptides for Increased Protein a Binding and/or Decreased Complement Binding and/or Decreased CD16 Binding
[0410] Purification of antibodies using Protein A affinity is a well-developed process. However, among four subtypes of canine IgG, only IgG-B Fc (e.g., SEQ ID NO: 2 or SEQ ID NO: 107) has Protein A binding affinity. Canine IgG-A Fc (e.g., SEQ ID NO: 1), IgG-C Fc (e.g., SEQ ID NO: 3 or SEQ ID NO: 108), and IgG-D Fc (e.g., SEQ ID NO: 4) have weak or no measurable Protein A binding affinity. Variant canine IgG-A Fc, IgG-C Fc, and IgG-D Fc polypeptides were designed for altered Protein A binding.
[0411] In addition, canine IgG-B Fc and IgG-C Fc have complement activity and bind to C1q, while canine IgG-A Fc and IgG-D Fc have weak or no measurable binding affinity to C1q. To potentially reduce the C1q binding and/or potentially reduce complement-mediated immune responses, variant canine IgG-B Fc and IgG-C Fc polypeptides were designed.
[0412] Furthermore, canine IgG-B Fc and IgG-C Fc have CD16 binding activity. To potentially reduce the binding of CD16 to IgG-B Fc and IgG-C Fc, and/or potentially reduce ADCC, variant canine IgG-B Fc and IgG-C Fc polypeptides were designed.
[0413] Table 3, below summarizes the Protein A and C1q binding characteristics of canine IgG Fc subtypes. Notably, none of the wild-type canine IgG Fc subtypes lacks C1q binding and binds Protein A.
TABLE-US-00003 TABLE 3 Wild-type Protein A C1q CD16 Canine IgG Fc Binding Binding Binding IgG-A Fc - - - IgG-B Fc + + + IgG-C Fc - + + IgG-D Fc - - - (-) denotes low or no measurable binding activity.
[0414] Using three-dimensional protein modeling and protein sequence analysis, the sequences of canine IgG-B Fc that are likely in contact with Protein A were identified. FIG. 1 shows an alignment of canine IgG-A, B, C, and D Fc sequences. The boxes indicate the regions likely in contact with Protein A.
[0415] Two approaches were used to design variant canine IgG-A, IgG-C, and IgG-D Fc polypeptides for increased Protein A binding. For the first approach, variant canine IgG-A, IgG-C, and IgG-D Fc polypeptides were designed to have the same Protein A binding motif sequences as canine IgG-B Fc (e.g., SEQ ID NO: 5, SEQ ID NO: 6, and SEQ ID NO: 7, respectively). For the second approach, variant canine IgG-A Fc I(21)T/Q(207)H (SEQ ID NO: 60), variant canine IgG-C Fc I(21)T (SEQ ID NO: 61), and variant canine IgG-D Fc I(21)T/Q(207)H (SEQ ID NO: 62) were designed with one or two amino acid substitutions in the Protein A binding region to correspond with the canine IgG-B Fc sequence.
[0416] In addition, variant canine IgG-A Fc, IgG-C Fc, and IgG-D Fc polypeptides with increased Protein A binding may be prepared having one or more of the amino acid substitutions listed in Table 4.
TABLE-US-00004 TABLE 4 Variant Canine IgG Fc Amino Acid Substitutions* (Protein A +) Canine IgG-A Fc Canine IgG-C Fc Canine IgG-D Fc (SEQ ID NO: 1) (SEQ ID NO: 3) (SEQ ID NO: 4) Ile (21) Thr Ile (21) Thr Ile (23) Thr Arg (23) Leu Val (23) Leu Arg (23) Leu Thr (25) Ala Thr (24) Ile Thr (25) Ala Glu (80) Gly Glu (80) Gly Thr (205) Ala Gln (207) His Gln (207) His *The amino acid positions listed are relative to the SEQ ID NO. indicated.
[0417] To potentially reduce the binding of C1q to canine IgG-B Fc and IgG-C Fc, and/or potentially reduce complement-mediated immune responses, variant canine IgG-B Fc and IgG-C Fc polypeptides may be prepared having an amino acid substitution of Lys with any amino acid except Lys at an amino acid position corresponding to position 93 of SEQ ID NO: 2 or of SEQ ID NO: 3, respectively. These amino acid substitutions were identified after analysis of the protein sequence and 3-D structure modeling of canine IgG-B Fc and IgG-C Fc compared to canine IgG-A Fc and IgG-D Fc, which are understood to not exhibit complement activity. For example, variant canine IgG-B Fc K(93)R (SEQ ID NO: 78) and variant canine IgG-C Fc K(93)R (SEQ ID NO: 79) may be prepared. Reduced binding between human C1q and a fusion protein comprising variant canine IgG-B Fc K(93)R was observed when compared to a fusion protein comprising wild-type canine IgG-B Fc.
[0418] To potentially reduce the binding of CD16 to IgG-B Fc and IgG-C Fc, and/or potentially reduce ADCC, variant canine IgG-B Fc and IgG-C Fc polypeptides may be prepared having one or more of the amino acid substitutions listed in Table 5. The amino acid substitution(s) were identified after analysis of the protein sequence and 3-D structure modeling of canine IgG-B and IgG-C compared to IgG-A and IgG-D, which are understood to not exhibit ADCC activity.
TABLE-US-00005 TABLE 5 Original residue position* Canine IgG-B Fc Canine IgG-C Fc (SEQ ID NO: 2) (SEQ ID NO: 3) Substitution(s) Met (5) Leu (5) Any amino acid except original residue, such as Pro Asp (38) Asp (38) Any amino acid except original residue, such as Gly Pro (39) Pro (39) Any amino acid except original residue, such as Arg Lys (97) Lys (97) Any amino acid except original residue, such as Ile Ala (98) Ala (98) Any amino acid except original residue, such as Gly *The amino acid positions listed are relative to the SEQ ID NO. indicated.
[0419] Since wild-type canine IgG-C Fc lacks Protein A binding and has C1q binding, a double variant canine IgG-C Fc that binds Protein A and has reduced binding to C1q may be prepared by combining one or more of the amino acid substitutions listed in Table 4 with a K(93)R substitution or K(93)X substitution, wherein X is any amino acid except Lys. A double variant canine IgG-B Fc or double variant canine IgG-C Fc with reduced binding to C1q and reduced binding to CD16 may be prepared by combining one or more of the amino acid substitutions listed in Table 5 with a K(93)R substitution or K(93)X substitution, wherein X is any amino acid except Lys. A triple variant canine-IgG-C Fc that binds Protein A and has reduced binding to C1q and CD16 may be prepared by combining one or more of the amino acid substitutions listed in Table 4 and one or more of the amino acid substitutions listed in Table 5 with a K(93)R substitution or K(93)X substitution, wherein X is any amino acid except Lys.
[0420] The binding of any variant canine IgG Fc to Protein A, CD16, and/or C1q may be determined and compared to the binding of another IgG Fc to Protein A, CD16, and/or C1q (e.g., the corresponding wild-type canine IgG Fc, another wild-type or variant canine IgG Fc, or a wild-type or variant IgG Fc of another companion animal, etc.).
[0421] Binding analysis may be performed using an Octet biosensor. Briefly, the target molecule (e.g., Protein A, C1q, CD16, etc.) may be biotinylated and free unreacted biotin removed (e.g., by dialysis). The biotinylated target molecule is captured on streptavidin sensor tips. Association of the target molecule with various concentrations (e.g., 10 .mu.g/mL) of IgG Fc polypeptide is monitored for a specified time or until steady state is reached. Dissociation is monitored for a specified time or until steady state is reached. A buffer only blank curve may be subtracted to correct for any drift. The data are fit to a 1:1 binding model using ForteBio.TM. data analysis software to determine the k.sub.on, k.sub.off, and the K.sub.d.
Example 2
Variant Equine IgG Fc Polypeptides for Increased Protein a Binding and/or Decreased Complement Binding
[0422] Of the seven subtypes of equine IgG, IgG1 Fc (e.g., SEQ ID NO: 63), IgG3 Fc (e.g., SEQ ID NO: 65), IgG4 Fc (e.g., SEQ ID NO: 66), IgG7 Fc (e.g., SEQ ID NO: 69) have Protein A binding affinity. Equine IgG2 Fc (e.g., SEQ ID NO: 18, SEQ ID NO: 64), IgG5 Fc (e.g., SEQ ID NO: 67), and IgG6 Fc (e.g., SEQ ID NO: 68) have weak or no measurable Protein A binding affinity. Variant equine IgG2 Fc, IgG5 Fc, and IgG6 Fc polypeptides were designed for altered Protein A binding.
[0423] In addition, equine IgG2 Fc, IgG5 Fc, and IgG6 Fc have weak or no measurable binding affinity to C1q, while equine IgG1 Fc, IgG3 Fc, IgG4 Fc, and IgG7 Fc bind to C1q. To potentially reduce the C1q binding and/or potentially reduce complement-mediated immune responses, variant equine IgG1 Fc, IgG3 Fc, IgG4 Fc, and IgG7 Fc polypeptides were designed.
[0424] Table 6, below summarizes the Protein A and C1q binding characteristics of equine IgG Fc subtypes. Notably, none of the wild-type equine IgG Fc subtypes lacks C1q binding and binds Protein A.
TABLE-US-00006 TABLE 6 Wild-type Protein A C1q Equine IgG Fc Binding Binding IgG1 Fc + + IgG2 Fc - - IgG3 Fc + + IgG4 Fc + + IgG5 Fc - - IgG6 Fc - - IgG7 Fc + + (-) denotes low or no measurable binding activity.
[0425] Using three-dimensional protein modeling and protein sequence analysis, the sequences of equine IgG1 Fc, IgG3 Fc, IgG4 Fc, and IgG7 Fc that are likely in contact with Protein A were identified. Variant equine IgG2 Fc, IgG5 Fc, and IgG6 Fc polypeptides with increased Protein A binding may be prepared having one or more of the amino acid substitutions listed in Table 7.
TABLE-US-00007 TABLE 7 Variant Equine IgG Fc Amino Acid Substitutions* (Protein A +) Equine IgG2 Fc Equine IgG5 Fc Equine Ig6 Fc (SEQ ID NO: 64) (SEQ ID NO: 67) (SEQ ID NO: 68) Ala (15) Thr Val (199) Leu Ile (199) Leu Phe (203) Tyr Glu (200) Tyr Arg (200) His His (201) Asn Thr (202) His *The amino acid positions listed are relative to the SEQ ID NO. indicated
[0426] For example, variant equine IgG2 Fc, IgG5 Fc, and IgG6 Fc polypeptides were designed with one or multiple amino acid substitutions in the Protein A binding region to correspond with the sequence of wild-type equine IgG Fc, which does bind Protein A. Variant equine IgG2 Fc F(203)Y (SEQ ID NO: 71); variant equine IgG2 Fc A(15)T/F(203)Y (SEQ ID NO: 72); variant equine IgG5 Fc V(199)L/E(200)Y (SEQ ID NO: 75); and variant equine IgG6 Fc I(199)L/R(200)H/H(201)N/T(202)H (SEQ ID NO: 76) with increased Protein A binding may be prepared.
[0427] To potentially reduce the binding of C1q to equine IgG1 Fc, IgG3 Fc, IgG4 Fc, and IgG7 Fc, and/or potentially reduce complement-mediated immune responses, variant canine IgG1 Fc, IgG3 Fc, IgG4 Fc, and IgG7 Fc polypeptides may be prepared having an amino acid substitution of Lys with any amino acid except Lys at an amino acid position corresponding to position 87 of SEQ ID NO: 63, of SEQ ID NO: 65, of SEQ ID NO: 66, of SEQ ID NO: 69, respectively. These amino acid substitutions were identified after analysis of the protein sequence and 3-D structure modeling of equine IgG1 Fc, IgG3 Fc, IgG4 Fc, and IgG7 Fc compared to equine IgG2 Fc, IgG5 Fc, and IgG6 Fc, which are understood to not exhibit complement activity. For example, variant equine IgG1 Fc K(87)S (SEQ ID NO: 70), variant equine IgG 3 Fc K(87)S (SEQ ID NO: 73), variant equine IgG4 Fc K(87)S (SEQ ID NO: 74), and variant equine IgG7 Fc K(87)S (SEQ ID NO: 77) may be prepared.
[0428] The binding of any variant equine IgG Fc to Protein A and/or C1q may be determined and compared to the binding of another IgG Fc to Protein A and/or C1q (e.g., the corresponding wild-type equine IgG Fc, another wild-type or variant equine IgG Fc, or a wild-type or variant IgG Fc of another companion animal, etc.). The binding assay described in Example 1 may be used.
Example 3
Variant Feline IgG Fc Polypeptides for Decreased Complement Binding
[0429] Each of the three subtypes of feline IgG, IgG1a Fc (SEQ ID NO: 80 or SEQ ID NO: 117), IgG1b Fc (SEQ ID NO: 81 or SEQ ID NO: 118), and IgG2 Fc (SEQ ID NO: 16) have Protein A binding affinity. However, only feline IgG2 Fc has weak or no measurable binding affinity to C1q, while feline IgG1a Fc, IgG1b Fc bind to C1q. To potentially reduce the C1q binding and/or potentially reduce complement-mediated immune responses, variant feline IgG1a Fc and IgG1b Fc polypeptides were designed.
[0430] Table 8, below summarizes the Protein A and C1q binding characteristics of feline IgG Fc subtypes. Notably, none of the wild-type equine IgG Fc subtypes lacks C1q binding and binds Protein A.
TABLE-US-00008 TABLE 8 Wild-type Protein A C1q Feline IgG Fc Binding Binding IgG1a Fc + + IgG1b Fc + + IgG2 Fc + - (-) denotes low or no measurable binding activity.
[0431] To potentially reduce the binding of C1q to feline IgG1a Fc and IgG1b Fc, and/or potentially reduce complement-mediated immune responses, variant feline IgG1a Fc and IgG1b Fc polypeptides may be prepared having an amino acid substitution of Pro with any amino acid except Pro at an amino acid position corresponding to position 198 of SEQ ID NO: 80 or of SEQ ID NO: 81, respectively. These amino acid substitutions were identified after analysis of the protein sequence and 3-D structure modeling of feline IgG1a Fc and IgG1b Fc compared to feline IgG2 Fc, which is understood to not exhibit complement activity. For example, variant feline IgG1a Fc P(198)A (SEQ ID NO: 82) and variant feline IgG1b Fc P(198)A (SEQ ID NO: 83) may be prepared.
[0432] The binding of any variant feline IgG Fc to C1q may be determined and compared to the binding of another IgG Fc to C1q (e.g., the corresponding wild-type feline IgG Fc, another wild-type or variant feline IgG Fc, or a wild-type or variant IgG Fc of another companion animal, etc.). The binding assay described in Example 1 may be used.
Example 4
Variant Canine and Feline IgG Fc Polypeptides for Heterodimeric Proteins
[0433] To enable the preparation of a bispecific canine or feline antibody or a bifunctional canine or feline Fc fusion protein using a knob-in-hole heterodimerization approach, pairing of variant canine IgG Fc polypeptides and variant feline IgG Fc polypeptides was investigated.
[0434] An amino acid substitution of threonine at a position corresponding to position 138 of canine IgG-A (SEQ ID NO: 1), at a position corresponding to position 137 of canine IgG-B Fc (SEQ ID NO: 2), at a position corresponding to position 137 of canine IgG-C Fc (SEQ ID NO: 3), or at a position corresponding to position 138 of canine IgG-D Fc (SEQ ID NO: 4) to tyrosine (T138Y or T137Y) can be introduced to one Fc chain (heterodimer chain 1). Examples of amino acid sequences of variant canine IgG-A Fc, IgG-B Fc, IgG-C Fc, and IgG-D Fc heterodimer chain 1 are SEQ ID NO: 8, SEQ ID NO: 10, SEQ ID NO: 12, and SEQ ID NO: 14, respectively.
[0435] An amino acid substitution of tyrosine at a position corresponding to position 181 of canine IgG-A (SEQ ID NO: 1), at a position corresponding to position 180 of canine IgG-B Fc (SEQ ID NO: 2), at a position corresponding to position 180 of canine IgG-C Fc (SEQ ID NO: 3), or at a position corresponding to position 181 of canine IgG-D Fc (SEQ ID NO: 4) to threonine (Y181T or Y180T) can be introduced to a second Fc chain (heterodimer chain 2). Examples of amino acid sequences of variant canine IgG-A Fc, IgG-B Fc, IgG-C Fc, and IgG-D Fc heterodimer chain 2 are SEQ ID NO: 9, SEQ ID NO: 11, SEQ ID NO: 13, and SEQ ID NO: 15, respectively.
[0436] A second pairing of variant canine IgG Fc heterodimer chains 3 and 4 was also investigated. An amino acid substitution of threonine to tryptophan at a position corresponding to position 138 of canine IgG-A (SEQ ID NO: 1) or of canine IgG-D (SEQ ID NO: 4) (T138W), or at a position corresponding to position 137 of canine IgG-B Fc (SEQ ID NO: 2) or of canine IgG-C Fc (SEQ ID NO: 3) (T137W) can be introduced to one Fc chain (heterodimer chain 3). Examples of amino acid sequences of variant canine IgG-A Fc, IgG-B Fc, IgG-C Fc, and IgG-D Fc heterodimer chain 3 are SEQ ID NO: 109, SEQ ID NO: 111, SEQ ID NO: 113, and SEQ ID NO: 115.
[0437] An amino acid substitution of threonine to serine at a position corresponding to position 138, of leucine to alanine at a position corresponding to position 140, and of tyrosine to threonine at a position corresponding to position 181 of canine IgG-A (SEQ ID NO: 1) or of IgG-D (SEQ ID NO: 4) (T138S, L140A, Y181T), or of threonine to serine at a position corresponding to position 137, of leucine to alanine at a position corresponding to position 139, and of tyrosine to threonine at a position corresponding to position 180 of canine IgG-B Fc (SEQ ID NO: 2) or of IgG-C(SEQ ID NO: 3) (T137S, L139A, Y180T) can be introduced to a second Fc chain (heterodimer chain 4). Examples of amino acid sequences of variant canine IgG-A Fc, IgG-B Fc, IgG-C Fc, and IgG-D Fc heterodimer chain 4 are SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, and SEQ ID NO: 116.
[0438] An amino acid substitution of threonine to tryptophan at a position corresponding to position 154 of feline IgG2 (SEQ ID NO: 16), of feline IgG1a Fc (SEQ ID NO: 80 or SEQ ID NO: 117), or of feline IgG1b Fc (SEQ ID NO: 81 or SEQ ID NO: 118) (T154W) can be introduced to one Fc chain (heterodimer chain 1). Examples of amino acid sequences of variant feline IgG2 Fc, IgG1a Fc, and IgG1b Fc heterodimer chain 1 are SEQ ID NO: 119, SEQ ID NO: 121, and SEQ ID NO: 123, respectively.
[0439] An amino acid substitution of threonine to serine at a position corresponding to position 154, of leucine to alanine at a position corresponding to position 156, and of tyrosine to threonine at a position corresponding to position 197 of feline IgG2 Fc (SEQ ID NO: 16), of IgG-1a (SEQ ID NO: 80 or SEQ ID NO: 117), or of IgG-1b Fc (SEQ ID NO: 81 or SEQ ID NO: 118) (T154S, L156A, Y197T) can be introduced to a second Fc chain (heterodimer chain 4). Examples of amino acid sequences of variant feline IgG2 Fc, IgG1a Fc, and IgG1b Fc heterodimer chain 4 are SEQ ID NO: 120, SEQ ID NO: 122, and SEQ ID NO: 124.
[0440] The pairing of variant canine IgG Fc heterodimer chains 1 and 2, the pairing of variant canine IgG Fc heterodimer chains 3 and 4, or the pairing of variant feline IgG Fc heterodimer chains 1 and 2 may allow for Fc heterodimerization and prevent or reduce Fc homodimerization. A heterodimer chain 1 of one canine IgG subtype may be combined with a heterodimer chain 2 of the same or a different canine IgG subtype. A heterodimer chain 3 of one canine IgG subtype may be combined with a heterodimer chain 4 of the same or a different canine IgG subtype. A heterodimer chain 1 of one feline IgG subtype may be combined with a heterodimer chain 2 of the same or a different feline IgG subtype. The design can enable dimerization of bispecific canine or feline antibodies. In addition, two different peptides or proteins or a combination of different proteins can be fused to the heterodimeric Fc chains. For example, a dual GLP1 and glucagon molecule can be created using variant canine IgG Fc heterodimer chains or variant feline IgG Fc heterodimer chains, such as a GLP1 polypeptide (e.g., SEQ ID NO: SEQ ID NO: 85, SEQ ID NO: 86, SEQ ID NO: 87, SEQ ID NO: 88, SEQ ID NO: 89, SEQ ID NO: 90, or SEQ ID NO: 91) fused to a variant canine IgG-D Fc heterodimer chain 1 (e.g., SEQ ID NO: 14) and a glucagon polypeptide (e.g., SEQ ID NO: 92, SEQ ID NO: 93, SEQ ID NO: 94, or SEQ ID NO: 95) fused to a variant canine IgG-D Fc heterodimer chain 2 (e.g., SEQ ID NO: 15).
Example 5
GLP1 Fusion Proteins
[0441] The amino acid sequence of GLP1 is conserved among human, feline, canine, and equine, among other species. GLP1 proteolytic products (e.g., GLP1 (amino acids 7-37) (SEQ ID NO: 85) are rapidly degraded by dipeptidyl peptidase-4 (DPP-4) and are understood to have a short serum half-life of a couple minutes.
[0442] The design of contiguous polypeptides comprising at least one GLP1 polypeptide and feline, canine, or equine IgG Fc polypeptides was investigated to generate long-acting GLP1 fusion proteins. The following constructs were designed:
GLP1A-L1-Fc; Formula (I):
Fc-L1-GLP1A; or Formula (II):
GLP1A-L1-Fc-L2-GLP1B. Formula (III):
wherein GLP1A is a first GLP1 polypeptide, GLP1B is a second GLP1 polypeptide, L 1 and L2 are linkers; and Fc is an IgG Fc polypeptide of a companion animal species.
[0443] GLP1 was modified to be DPP-4 resistant by replacing alanine with either glycine or serine at a position corresponding to position 8 of wild-type GLP1 (7-37) (SEQ ID NO: 85). A minimal sequence of wild-type GLP1 (7-37) (SEQ ID NO:87) for binding to the N-terminal domain of mature feline GLP1R (SEQ ID NO: 49) was analyzed by three-dimensional protein modeling of the complex. Based on this modeling, the two C-terminal amino acids were removed from the DPP-4 resistant GLP1 to generate GLP1-S8 (7-35) (SEQ ID NO: 86) and GLP1-G8 (7-35) (SEQ ID NO: 87) polypeptides.
[0444] GLP1 polypeptides when positioned at the C-terminus of a construct, such as in formulas II and III, are not susceptible to DPP-4 degradation. Therefore, the alanine to glycine or serine substitution is not necessary for GLP1 polypeptides positioned at the C-terminus. Accordingly, wild-type GLP1 (7-37) (SEQ ID NO: 85) may be used at the C-terminus.
[0445] The linker may be a flexible, non-structural linker, such as a glycine- and serine-rich linker. A flexible extension may be added to the C-terminus of the contiguous polypeptide. The extension may comprise a glycine residue (SEQ ID NO: 88), two glycine residues (SEQ ID NO: 89), a three glycine residues (SEQ ID NO: 90), four glycine residues (SEQ ID NO: 91), five glycine residues (SEQ ID NO: 92), six glycine residues (SEQ ID NO: 93), seven glycine residues (SEQ ID NO: 94), eight glycine residues (SEQ ID NO: 95), or more glycine residues.
Example 6
GLP1 and Feline IgG Fc Fusion Proteins
[0446] Nucleotide sequences encoding (1) a contiguous polypeptide of Formula I having a signal sequence, GLP1-G8 (7-35) (SEQ ID NO: 87), a flexible linker, and wildtype feline IgG2 Fc (ssGLP1-G8_I_WTfeIgG2; SEQ ID NO: 96); and (2) a contiguous polypeptide of Formula III having a signal sequence, GLP1-G8 (7-35) (SEQ ID NO: 87), GLP1 (7-35) (SEQ ID NO: 89), two flexible linkers, a 2G C-terminal extension, and wildtype feline IgG2 Fc (ssGLP1-G8/GLP1-2G_III_WTfeIgG2; SEQ ID NO: 97) were synthesized and cloned into separate mammalian expression vectors. Wild-type feline IgG2 Fc (SEQ ID NO: 16) was chosen based on its low or no C1Q binding for reduced complement activity and Protein A binding for ease of purification.
[0447] The resulting vectors were separately transfected to CHO cells. The supernatant containing the contiguous polypeptides without the signal peptide (SEQ ID NOs: 24 and 23) was collected and filtered. Both proteins were affinity purified using a Protein A column (CaptivA.RTM. Protein A Affinity Resin, Repligen). The proteins were determined to be monomeric as assessed by HPLC gel filtration. However, the SDS-PAGE analysis showed that a percentage of both continuous polypeptides migrated to the same position in the gel in the absence and presence of reducing agent (DTT) (FIG. 2A). This result suggests that the hinge of wild-type feline IgG2 Fc, which has only one pair of cysteine residues, may not be enough to form an effective disulfide linkage.
[0448] Three-dimensional protein modeling analysis of several ortholog hinge structures was used to determine the approximate locations for modifying the feline IgG2 hinge to increase disulfide formation. To increase disulfide formation at the feline IgG2 hinge, the hinge sequence may be modified by substituting an amino acid with cysteine. For example, a variant feline IgG2 Fc (SEQ ID NO: 17) having a modified hinge was prepared by substituting Gly with Cys at an amino acid position corresponding to position 14 of SEQ ID NO: 16. The corresponding contiguous polypeptides ssGLP1-G8_I_VARfeIgG2 (SEQ ID NO: 39) and ssGLP1-G8/GLP1-2G_III_WTfeIgG2 (SEQ ID NO: 38) comprising variant feline IgG Fc of SEQ ID NO: 17 were designed, expressed in CHO cells, and purified by Protein A chromatography. The amino acid sequences of the secreted proteins after cleavage of the signal sequence are SEQ ID NOs 26 and 25, respectively. The SDS-PAGE analysis of the variant feline IgG2 constructs showed a decrease in the amount of protein in the lower molecular weight band in absence of reducing agent compared to the wild-type feline IgG2 constructs (compare FIG. 2 B to FIG. 2A). These results suggest that the Fc covalent pairing was improved for both variant feline IgG2 constructs.
[0449] Furthermore, differential scanning fluorimetry was used to assess the stability of the contiguous polypeptides at various pH, as reflected by mean melting point temperature (n=3) (Table 9, below). The increased stability of the variant feline IgG2 hinge is most evident at pH 6. For example, the constructs having variant feline IgG2 (SEQ ID NOs: 25 and 26) exhibited a higher Tm at pH 6 (56.9 and 59.7.degree. C.) than the corresponding constructs having wild-type feline IgG2 (SEQ ID NOs: 23 and 24), which had a Tm of 55.2 and 56.9.degree. C., respectively.
TABLE-US-00009 TABLE 9 Mean Melting Point Temperature (Tm .degree. C.) (n = 3) Construct (10 .mu.g) pH 3 pH 4 pH 5 pH 6 pH 7 pH 8 GLP1-G8/ NC NC NC 55.2 55.7 54.2 GLP1-2G_III_WTfeIgG2 (SEQ ID NO: 23) GLP1-G8_I_WTfeIgG2 NC NC 48.5 56.9 59.9 59 (SEQ ID NO: 24) GLP1-G8/ NC NC NC 56.9 55 52.5 GLP1-2G_III_WTfeIgG2 (SEQ ID NO: 25) GLP1-G8_I_VARfeIgG2 NC NC 53.1 59.7 59.9 58.2 (SEQ ID NO: 26) NC = no curve because no distinct transition point was observed.
[0450] Contiguous polypeptides of Formulas I, II, and III comprising GLP1-S8 (7-35) (SEQ ID NO: 86) instead of GLP1-G8 (7-35) (SEQ ID NO: 87) may be similarly designed and prepared. For example, ssGLP1-S8_I_WTfeIgG2 (SEQ ID NO: 40), GLP1-S8_I_WTfeIgG2 (SEQ ID NO: 28), ssGLP1-S8/GLP1-3G_III_WTfeIgG2 (SEQ ID NO: 37), and GLP1-S8/GLP1-3G_III_WTfeIgG2 (SEQ ID NO: 27) may be prepared. Similar constructs having variant instead of wild-type feline IgG Fc, such as GLP1-S8_I_VARfeIgG2, GLP1-S8_VARfeIgG2, GLP1-S8/GLP1-2G_III_VARfeIgG2, and GLP1-S8/GLP1-3G_III_VARfeIgG2, may also be prepared.
[0451] While feline IgG2 Fc was used in this example, contiguous polypeptides of Formulas I, II, and III comprising feline IgG1a Fc or IgG1b Fc instead of IgG2 may be designed and prepared. For example, similar contiguous polypeptides having wild-type feline IgG1a Fc, wild-type feline IgG1b Fc, variant feline IgG1a Fc, or variant feline IgG1b Fc (SEQ ID NOs: 80 to 83, respectively), may be designed and prepared.
Example 7
GLP1 and Canine IgG Fc Fusion Proteins
[0452] Various Formula I, II, and III contiguous polypeptides comprising a variant GLP1 and a canine IgG Fc may be designed and prepared. For example, a variant canine IgGD Fc (e.g., SEQ ID NO: 7 or SEQ ID NO: 62) may be chosen based on its low or no C1q binding for reduced complement activity and Protein A binding for ease of purification. In addition, a flexible, non-structural linker, such as a glycine- and/or serine-rich linker, may be used.
[0453] GLP1-G8_I_VARcaIgGD (SEQ ID NO: 30) and GLP1-S8_I_VARcaIgGD (SEQ ID NO: 32) are examples of Formula I contiguous polypeptides comprising (1) either GLP1-G8 (7-35) or GLP1-S8 (7-35), (2) a flexible linker, and (3) a variant canine IgGD Fc (e.g., SEQ ID NO: 7). GLP1-G8/GLP1-3G_III_VARcaIgGD (SEQ ID NO: 29) and GLP1-S8/GLP1-3G_III_VARcaIgGD (SEQ ID NO: 31) are examples of Formula III contiguous polypeptides comprising (1) either GLP1-G8 (SEQ ID NO: 87) or GLP1-S8 (SEQ ID NO: 86), (2) GLP1 (7-35) (SEQ ID NO: 61), (3) two flexible linkers, (4) a 3G C-terminal extension, and (5) a variant canine IgGD Fc.
[0454] The contiguous polypeptides may be designed with a signal sequence, a different GLP1 polypeptide (e.g., SEQ ID NO: 85, 86, 87, 98, or 99), or different modifications to the canine IgG Fc. Examples with such variations include ssGLP1-S8/GLP1-2G_III_VARcaIgGD (SEQ ID NO: 41), ssGLP1-G8/GLP1-2G_III_VARcaIgGD (SEQ ID NO: 42), ssGLP1-S8/GLP1-3G_III_VARcaIgGD (SEQ ID NO: 43), GLP1-S8/GLP1-2G_III_VARcaIgGD (SEQ ID NO: 105), and GLP1-G8/GLP1-2G_III_VARcaIgGD (SEQ ID NO: 106).
[0455] Furthermore, contiguous polypeptides of Formulas I, II, and III may comprise a wild-type canine IgGD, or a wild-type or variant canine IgGA, IgGB, or IgGC, instead of a variant canine IgGD. For example, similar contiguous polypeptides may be designed and prepared having a wild-type canine IgGA Fc, IgGB Fc, IgGC Fc, or IgGD Fc (e.g., SEQ ID NO: 1, 2, 3, or 4, respectively). Additional examples include contiguous polypeptides comprising a variant canine IgGA Fc (e.g., SEQ ID NO: 5 or 60), a variant canine IgGB Fc (e.g., SEQ ID NO: 78), or a variant canine IgGC Fc (e.g., SEQ ID NO: 6, 61, 79, or 84).
[0456] GLP1-G8/GLP1-3G_III_VARcaIgGD (SEQ ID NO: 29) and GLP1-G8_I_VARcaIgGD (SEQ ID NO: 30) were expressed separately in CHO cells and the supernatants containing the proteins collected and filtered. Both contiguous polypeptides were affinity purified by Protein A chromatography. The SDS-PAGE profiles of the two contiguous polypeptides in the absence and presence of reducing agent (DTT) were compared and the results suggested that the Fc disulfide bond was effectively formed for both polypeptides (data not shown).
Example 8
GLP1 and Equine IgG Fc Fusion Proteins
[0457] Various Formula I, II, and III contiguous polypeptides comprising a variant GLP1 and an equine IgG Fc may be designed and prepared. For example, a variant canine IgG2 Fc (e.g., SEQ ID NO: 19, 71, or 72) may be chosen based on its low or no C1q binding for reduced complement activity and Protein A binding for ease of purification. In addition, a flexible, non-structural linker, such as a glycine- and/or serine-rich linker, may be used.
[0458] GLP1-G8_I_VAReqIgG2 (SEQ ID NO: 34) and GLP1-S8_I_VAReqIgG2 (SEQ ID NO: 36) are examples of Formula I contiguous polypeptides comprising (1) either GLP1-G8 or GLP1-S8, (2) a flexible linker, and (3) a variant equine IgG2 Fc (e.g., SEQ ID NO: 19). GLP1-G8/GLP1-3G_III_VAReqIgG2 (SEQ ID NO: 33) and GLP1-S8/GLP1-3G_III_VAReqIgG2 (SEQ ID NO: 35) are examples of Formula III contiguous polypeptides comprising (1) either GLP1-G8 (7-35) (SEQ ID NO: 87) or GLP1-S8 (7-35) (SEQ ID NO: 86), (2) GLP1 (7-35) (SEQ ID NO: 61), (3) two flexible linkers, (4) a 3G C-terminal extension, and (5) a variant equine IgGD Fc.
[0459] The contiguous polypeptides may be designed with a signal sequence, a different GLP1 analog (e.g., SEQ ID NO: 86, 87, 98, or 99), a glycine extension (e.g., SEQ ID NO: 88 to 95) or additional modifications to the equine IgG Fc. Examples with such variations include ssGLP1-G8/GLP1-3G_III_VAReqIgG2 (SEQ ID NO: 44), ssGLP1-G8_I_VAReqIgG2 (SEQ ID NO: 45), ssGLP1-S8/GLP1-3G_III_VAReqIgG2 (SEQ ID NO: 46), and ssGLP1-S8_I_VAReqIgG2 (SEQ ID NO: 47).
[0460] Furthermore, contiguous polypeptides of Formulas I, II, and III may comprise a wild-type equine IgG2, or a wild-type or variant equine IgG1, IgG3, IgG4, IgG5, IgG6, IgG7, instead of a variant equine IgG2. For example, similar contiguous polypeptides may be designed and prepared having a wild-type equine IgG1 Fc (e.g., SEQ ID NO: 63), IgG2 Fc (e.g., SEQ ID NO: 18 or 64), IgG3 Fc (e.g., SEQ ID NO: 65), IgG4 Fc (e.g., SEQ ID NO: 66), IgG5 Fc (e.g., SEQ ID NO: 67), IgG6 Fc (e.g., SEQ ID NO: 68), or IgG7 Fc (e.g., SEQ ID NO: 69). Additional examples include contiguous polypeptides comprising a variant equine IgG1Fc (e.g., SEQ ID NO: 70), a variant equine IgG3 Fc (e.g., SEQ ID NO: 73), a variant equine IgG4 Fc (e.g., SEQ ID NO: 74), a variant equine IgG5 Fc (e.g., SEQ ID NO: 75), a variant equine IgG6 Fc (e.g., SEQ ID NO: 76), or a variant equine IgG7 Gc (e.g., SEQ ID NO: 77).
[0461] The nucleotide sequences encoding ssGLP1-G8/GLP1-3G_III_VAReqIgG2 (SEQ ID NO: 44) and ssGLP1-G8_I_VAReqIgG2 (SEQ ID NO: 45) were synthesized and cloned into separate mammalian expression vectors. The resulting vectors were separately transfected to CHO cells. The supernatant containing the contiguous polypeptides following cleavage of the signal peptide (SEQ ID NOs: 103 and 104) was collected and filtered. Both proteins were affinity purified using a Protein A column (CaptivA.RTM. Protein A Affinity Resin, Repligen). were expressed separately in CHO cells and the supernatants containing the proteins collected and filtered. Both contiguous polypeptides were affinity purified by Protein A chromatography. The SDS-PAGE profiles of the two contiguous polypeptides in the absence and presence of reducing agent (DTT) were compared and the results suggested that the Fc disulfide bond was effectively formed for both polypeptides (data not shown).
Example 9
Expression and Purification of Feline GLP1R N-Terminal Soluble Domain
[0462] The N-terminal domain of mature feline GLP1R (SEQ ID NO: 49) responsible for binding GLP1 to GLP1R was identified from the full-length feline GLP1R amino acid sequence (SEQ ID NO: 48). Nucleotide sequences encoding (1) a signal sequence, feline N-terminal GLP1R, human Fc, and a poly-His tag (ssFeGLP1R-N-huFc_PolyHis; SEQ ID NO: 50) and (2) a signal sequence, feline N-terminal GLP1R, and a poly-His tag (ssFeGLP1R-N_polyHis; SEQ ID NO: 51) were synthesized and cloned into separate mammalian expression vectors. The resulting vectors were separately transfected to CHO cells. The supernatant containing the polypeptides was collected and filtered. The proteins were affinity purified using a Ni-Sepharose column or Protein A column (CaptivA.RTM. Protein A Affinity Resin, Repligen) for the huFc construct. Both proteins were used for GLP1 functional binding and ELISAs.
Example 10
GLP1 Fusion Protein Binding Kinetics
[0463] The binding affinity of GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) to FeGLP1R-N-huFc_PolyHis was assessed using biolayer interferometry (Octet). Briefly, FeGLP1R-N-huFc_PolyHis was biotinylated, the free unreacted biotin was removed, and the biotinylated protein was captured to streptavidin sensor tips. The association of different concentrations GLP1-G8_I_VARfeIgG2 was monitored for ninety seconds. Dissociation was monitored for 600 seconds. A buffer only blank curve was subtracted to correct for any drift. The data were fit to a 1:1 binding model using ForteBio.TM. data analysis software to determine the k.sub.on, k.sub.off, and the K.sub.d. The buffer for dilutions and all binding steps was: 20 mM phosphate, 150 mM NaCl, pH 7.2. The K.sub.d between Feline GLP1RN-terminal domain and GLP1-G8_I_VARfeIgG2 was between 8.0.times.10.sup.-9 and 16.times.10.sup.-9 M.
Example 11
GLP1 Fusion Protein Bioactivity in a Cell-Based Assay
[0464] CHOK-1-GL1R cell line (Discoverx, cat #95-0062C2), a hamster ovarian cell line which overexpresses Gs-coupled human glucagon ligand peptide-1 receptors (GLP1R) on the cell surface, was used to measure GLP1 cellular activity with a cAMP Hunter Bioassay kit (Discoverx, Cat #95-0062Y2). Cells were plated in a 96-well plate and incubated at 37.degree. C., 5% CO.sub.2 for 24 hours. Cells were then treated with a control agonist--either GLP1 human (37 a.a.) (Prospec, Cat #HOR-236) or Extendin-4 (Discoverx, Cat #92-1115)--or a contiguous polypeptide comprising a variant GLP1 at a series of 3-fold dilutions followed by incubation at 37.degree. C. for 30 min. The contiguous polypeptides tested were GLP1-G8/GLP1-2G_III_VARfeIgG2 (SEQ ID NO: 25) and GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26).
[0465] Upon GLP1 binding to the Gs-coupled GLP1R receptor, Gs stimulates adenylate cyclase to generate cAMP. At the end of incubation, cAMP Antibody Reagent and cAMP Working Detection Solution, which contains lysis buffer, .beta.-galactosidase (.beta.-gal) small fragment conjugated cAMPs, and substrates, were added to the cells. The cells were incubated in the dark for 1 hour at room temperature to allow the immunocompetition reaction to occur between endogenously generated cAMPs and .beta.-gal small fragment conjugated cAMPs for cAMP antibody binding.
[0466] At the end of the 1 hour incubation, cAMP Solution A containing .beta.-gal large fragments, which can complement with the free (non-antibody binding) .beta.-gal small fragment-cAMPs to form functional enzymes, were added to the cell lysate. The lysate was incubated for 3 to 6 hours in the dark at room temperature to allow the .beta.-gal to hydrolyze the substrate and generate luminescent signals.
[0467] The more unbound free .beta.-gal small fragment-cAMPs that remain, the more functional .beta.-gal enzymes form. Therefore, the amount of signal produced is directly proportional to the amount of cAMP in the cell lysate. At the end of incubation, luminescence was read on a Synergy HT microplate reader (Biotek, Winooski, Vt.). The EC50s were calculated with a software GraphPad Prism (GraphPad Software, Inc. La Jolla, USA).
[0468] FIG. 3 shows a plot of the relative light units (RLU) versus concentration for each sample tested. The EC50s are listed in Table 10, below. Both GLP1-G8/GLP1-2G_III_VARfeIgG2 (SEQ ID NO: 25) and GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) are more active than GLP1 (7-37) and Extendin-4. GLP1-G8/GLP1-2G_III_VARfeIgG2 (SEQ ID NO: 25) is more potent than GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26), suggesting that the C-terminal GLP1 may contribute additional activity.
TABLE-US-00010 TABLE 10 Sample EC 50 (nM) GLP1 (7-37) 0.1449 Extendin-4 0.4596 GLP1-G8/GLP1-2G_III_VARfeIgG2 0.01751 (SEQ ID NO: 25) GLP1-G8_I_VARfeIgG2 0.02806 (SEQ ID NO: 26)
Example 12
GLP1 Fusion Protein Long-Term Stability
[0469] GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) was stored in PBS, pH7.2 at a concentration of 1.3 mg/mL, placed in 1.5 mL Eppendorf tube, and stored at 2-8.degree. C. for one year (Lot 2-29-2016). GLP1-G8_IVARfeIgG2 (SEQ ID NO: 26) was also stored in PBS, pH7.2 at a concentration of 10 mg/mL, placed in 1.5 mL Eppendorf tube, and stored at 2-8.degree. C. for one day (Lot 2-2-2017). To evaluate stability, the stored sample were analyzed by cell-based assay using the same CHOK-1-GL1R cell line (Discoverx, cat #95-0062C2) described in Example 11, but cellular activity was assessed using a cAMP-Glo.TM. Max Assay (Promega, Cat #PAV1682).
[0470] CHOK-1-GL1R cells were plated in a 96-well plate (Corning, Cat #3610) at a density of 20,000 cells per well in F-12K Medium (ATCC, Cat #ATCC.RTM. 30-2004) supplemented with 10% Fetal Bovine Serum, heat inactivated (Sigma, Cat #2868) and incubated at 37.degree. C., 5% CO2 for 24 hours. Cells were then stimulated with a control agonist GLP1 human (37 a.a.) (Prospec, Cat #HOR-236), Lot 2-20-2016, or Lot 2-2-2017 at a series of 3-fold dilutions with serum-free medium followed by addition of Complete Induction Buffer which contains MgCl.sub.2 to a final concentration 20 mM, isobutyl-1-methylxanthine (IBMX) (Sigma-Aldrich Cat.#17018) to a final concentration 500 .mu.M and Ro 20-1724 [4-(3-butoxy-4-methoxy-benzyl) imidazolidone] (Sigma Aldrich, Cat.#B8279) to a final concentration 100 .mu.L.
[0471] The cells were incubated at room temperature for 30 minutes. In this process, upon GLP1 binding to the Gs-coupled GLP1R receptor, Gs stimulates adenylate cyclase to generate cAMP. At the end of incubation, cAMP Detection Solution, which contains an inactive protein kinase A holoenzyme, protein kinase A substrate, and lysis buffer, was added to the cells. The plates placed on an orbital shaker for 1-2 minutes and then incubated at room temperature (23.degree. C.) for 20 minutes. Cellular cAMP will activate protein kinase A by binding its regulatory-inhibitory subunits and releasing the catalytic subunits. The free catalytic subunits catalyze the transfer of the terminal phosphate of ATP to the protein kinase A substrate, consuming cellular ATP in the process. At the end of incubation, a luciferase-based Kinase-Glo.RTM. Reagent was added to the cell lysates and the plates were shaken on an orbital shaker for 2 min followed by incubation in the dark at room temperature for 10 min.
[0472] Mono-oxygenation of luciferin was catalyzed by luciferase in the presence of Mg.sup.2+ and ATP that presented in the cell lysate, resulting in a luminescent signal proportional to the amount of ATP in the cells. At the end of 10 min incubation, the plate was read on a Synergy HT microplate reader (Biotek, Winooski, Vt.). Luminescence is proportional to ATP levels but inversely proportional to cAMP levels. Thus, as cAMP concentration increases, luminescence decreases.
[0473] FIG. 4 shows the results of the cell-based assay as a plot of luminescence versus concentration for Lot 2-29-2016 compared to Lot 2-2-2017 and GLP1 (31 a.a.). The GLP1-G8_I_VARfeIgG2 sample maintained cellular activity after storage in PBS at 2-8.degree. C. for one year.
Example 13
GLP1 Fusion Protein Serum Stability
[0474] GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) was stored in PBS, pH7.2 with feline serum at 37.degree. C. for 24 hours to test in vitro serum stability. The cell-based assay was performed as described in Example 12 and results suggested that the activity was maintained (data not shown). In addition, no visible degradations were observed by Western blot analysis (FIG. 5).
Example 14
GLP1 Fusion Protein In Vivo Pharmacokinetics
[0475] GLP1-G8_I_VARfeIgG2 (SEQ ID NO: 26) was administered as a single dose (2 mg/kg) by subcutaneous injection to 5 cats. Serum samples were taken before dosing (time 0) and at 4 hours, 8 hours, 12 hours, 24 hours, 48 hours, 72 hours, and 168 hours. The concentration of GLP1-G8_I_VARfeIgG2 in the serum samples was measured by quantitative ELISA. GLP1-G8_I_VARfeIgG2 polypeptide with a detection limit of 4 ng/mL was used as a reference. The serum concentration of GLP1-G8_I_VARfeIgG2 was plotted against time (FIG. 6). The mean serum half-life (t.sub.1/2) of GLP1-G8_I_VARfeIgG2 was 39 hours. The average T.sub.max was 22 hours, the average C.sub.max was 12 .mu.g/mL, and the mean area under the curve (AUC) was about 950 m(h)/mL.
[0476] The quantitative ELISA used an anti-GLP1 antibody (4F3, Novus Biologicals, Catalog No. NBP1-97413) and a goat anti-cat IgG-Fc, HRP conjugated antibody (Bethyl Laboratories, Inc., Catalog No. A20-117P) for quantification of GLP1-G8_I_VARfeIgG2 in feline serum samples from the in vivo pharmacokinetics study. A 96-well plate was coated with anti-GLP1 antibody (5 .mu.g/mL in coating buffer, 100 .mu.l/well). The plate was sealed and incubated overnight at 4.degree. C. The plate was washed in triplicate with 1.times.TBST (10.times.TBST, Teknova, Catalog No. T9511) and blocking buffer was added. After removing the blocking buffer, serial dilutions of reference standard and samples in blocking buffer were added (100 .mu.l/well) and the plate was incubated for 2 hours at room temperature. The plate was washed in triplicate with 1.times.TBST and goat anti-cat IgG Fc antibody was added (0.1 .mu.g/mL in blocking buffer, 100 .mu.l/well). After incubation for 1 hour at room temperature, the plate was washed 5 times with 1.times.TBST. TMB substrate (ScyTek, Catalog No. TM1999) was added (100 .mu.l/well) and allowed to incubate at room temperature for 1 minute. The reaction was stopped by the addition of 2M H.sub.2SO.sub.4 (50 .mu.l/well). Absorbance at 450 nm was measured and the concentration of GLP1-G8_I_VARfeIgG2 in the serum samples calculated.
[0477] Furthermore, GLP1-G8_I_VARfeIgG2 concentration in the same serum samples (no DPP-4 inhibitor added) was assessed using a cell-based activity assay. The same CHOK-1-GL1R cell line (Discoverx, cat #95-0062C2) and cAMP-Glo.TM. Max Assay (Promega, Cat #PAV1682) described in Example 12 was used. In this Example, however, the cells were stimulated with a control agonist GLP1 human (37 a.a.) (Prospec, Cat #HOR-236) or samples of cat serum taken before dosing (time 0) and at at 4 hours, 8 hours, 12 hours, 24 hours, 48 hours, 72 hours, and 168 hours diluted in serum-free medium (5%, 0.5%, and 0.05% dilutions). The concentration of GLP1-G8_I_VARfeIgG2 in each sample was calculated using SoftMax pro 7 (Molecular Devices, Sunnyvale, Calif.). The mean concentration of GLP1-G8_I_VARfeIgG2 for the 5 cats was plotted against time (FIG. 7). The mean AUC was about 840 .mu.g(h)/mL and the mean t.sub.1/2 was 36 hours. The concentrations calculated from the cell-based activity assay are consistent with the concentrations obtained from the ELISA, which suggests that the variant GLP1 detected by ELISA is biologically active.
Example 15
GLP1, Glucagon, and IgG Fc Fusion Proteins
[0478] To investigate a long acting GLP1 receptor and Glucagon receptor dual agonist, contiguous polypeptides comprising a GLP1 polypeptide, a glucagon polypeptide, and an IgG Fc polypeptide having the following constructs were designed:
GLP1-L1-Fc-L2-Gluc; and Formula (IV):
Gluc-L1-Fc-L2-GLP1, Formula (V):
wherein GLP1 is a GLP1 polypeptide, Gluc is a glucagon polypeptide, L1 and L2 are linkers, and Fc is an Fc polypeptide.
[0479] As discussed above, GLP1 was modified to be DPP-4 resistant by replacing alanine with either glycine or serine at a position corresponding to position 8 of wild-type GLP1 (7-37) (SEQ ID NO: 85). In addition, the DPP-4 resistant GLP1 was further modified by removing the two C-terminal amino acids to generate variant GLP1-S8 (7-35) (SEQ ID NO: 86) and variant GLP1-G8 (7-35) (SEQ ID NO: 87) polypeptides.
[0480] GLP1 polypeptides when positioned at the C-terminus of a construct, such as in Formula (V), are not susceptible to DPP-4 degradation. Therefore, the alanine to glycine or serine substitution is not necessary for GLP1 polypeptides positioned at the C-terminus. Accordingly, wild-type GLP1 (7-37) (SEQ ID NO: 85) may be used at the C-terminus.
[0481] The linker may be a flexible, non-structural linker, such as a glycine- and serine-rich linker. A flexible extension may be added to the C-terminus of the contiguous polypeptide. The extension may comprise a glycine residue (SEQ ID NO: 88), two glycine residues (SEQ ID NO: 89), a three glycine residues (SEQ ID NO: 90), four glycine residues (SEQ ID NO: 91), five glycine residues (SEQ ID NO: 92), six glycine residues (SEQ ID NO: 93), seven glycine residues (SEQ ID NO: 94), eight glycine residues (SEQ ID NO: 95), or more glycine residues.
[0482] The contiguous polypeptide may comprise a wildtype glucagon polypeptide (e.g., SEQ ID NO: 21) or a variant glucagon polypeptide.
[0483] The contiguous polypeptide may comprise a human IgG Fc or an IgG Fc of a companion animal species, such as canine, feline, or equine. The subtype of IgG Fc used may be based on having low or no C1q binding activity and/or having Protein A binding capacity. For example, a wild-type or variant human, canine, equine, or feline IgG Fc having low or no C1q binding and/or having Protein A binding capacity may be used (e.g., SEQ ID NO: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, or 84).
[0484] Exemplary contiguous polypeptides comprising a GLP1 polypeptide, a Glucagon polypeptide, and a feline IgG Fc polypeptide include GLP1-G8/Gluc-3G_IV_WTfeIgG2 (SEQ ID NO: 52) and Gluc/GLP1-2G_V_WTfeIgG2 (SEQ ID NO: 53).
[0485] Exemplary contiguous polypeptides comprising a GLP1 polypeptide, a Glucagon polypeptide, and a canine IgG Fc polypeptide include GLP1-G8/Gluc-4G_IV_VARcaIgGD (SEQ ID NO: 54) and Gluc/GLP1-3G_V_VARcaIgGD (SEQ ID NO: 55).
[0486] Exemplary contiguous polypeptides comprising a GLP1 polypeptide, a Glucagon polypeptide, and an equine IgG Fc polypeptide include GLP1-G8/Gluc-4G_IV_VAReqIgGD (SEQ ID NO: 56) and Gluc/GLP1-3G_V_VAReqIgG2 (SEQ ID NO: 57).
[0487] Exemplary contiguous polypeptides comprising a GLP1 polypeptide, a Glucagon polypeptide, and a human IgG Fc polypeptide include GLP1-G8/Gluc-4G_IV_huIgG4 (SEQ ID NO: 8) and Gluc/GLP1-3G_V_huIgG4 (SEQ ID NO: 59).
Example 16
Variant IgG Fc Polypeptides for Enhanced Hinge Disulfide Formation
[0488] Additional three-dimensional protein modeling analysis of several ortholog hinge structures was used to modify feline and equine IgG hinges to enhance disulfide formation. To enhance disulfide formation at the feline IgG hinge, the hinge sequence may be modified by substituting lysine with proline at a position corresponding to position 16 of feline IgG2 (SEQ ID NO: 16), of feline IgG1a (SEQ ID NO: 80 or SEQ ID NO: 117), or of feline IgG1b (SEQ ID NO: 81 or SEQ ID NO: 118) (e.g., K16P). Examples of amino acid sequences of variant feline IgG polypeptides having a modified hinge include SEQ ID NO: 125, SEQ ID NO: 126, and SEQ ID NO: 127.
[0489] To enhance disulfide formation at the equine IgG hinge, the hinge sequence may be modified by substitution cysteine with serine at a position corresponding to position 3 of an equine IgG (e.g., IgG2 Fc (SEQ ID NO: 129)) and/or substituting glutamine with proline at a position corresponding to position 20 of an equine IgG (e.g., IgG2 Fc (SEQ ID NO: 129) (e.g., C3S, Q20P). Examples of amino acid sequences of variant equine IgG polypeptides having a modified hinge include SEQ ID NO: 130, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 134, and SEQ ID NO: 135.
Example 17
Variant IgG Fc Polypeptides for Enhanced Recombinant Production and/or Enhanced Hinge Disulfide Formation
[0490] Three-dimensional protein modeling was used to design feline and equine variant IgG Fc polypeptides comprising sequences from the hinge region from a different IgG isotype for enhanced recombinant production and improved hinge disulfide formation. Variant feline IgG2 Fc polypeptides may be prepared that comprise sequences from the hinge region of feline IgG1a or IgG1b (e.g., SEQ ID NO: 125). In addition, variant equine IgG2 Fc polypeptides may be prepared that comprise sequences from the hinge region of equine IgG1 (e.g., SEQ ID NO: 19).
[0491] Levels of recombinant production of variant IgG Fc polypeptides and/or levels of hinge disculfide formation may be determined and compared to that of another IgG Fc by SDS-PAGE analysis under reducing and non-reducing conditions (e.g., the corresponding wild-type IgG Fc of the same or different isotype, or a wild-type or variant IgG Fc of another companion animal, etc.).
Sequence CWU
1
1
1991221PRTUnknownWild-type canine IgG-A Fc , Protein A -, C1q -, CD16
1Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val Leu Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Ile Leu
Arg Ile Thr Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln
Ile Ser Trp 35 40 45Phe Val Asp
Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg Glu 50
55 60Gln Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val
Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His
85 90 95Ile Asp Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Arg Ala His Lys Pro Ser Val Tyr Val Leu Pro Pro
Ser Pro Lys Glu 115 120 125Leu Ser
Ser Ser Asp Thr Val Ser Ile Thr Cys Leu Ile Lys Asp Phe 130
135 140Tyr Pro Pro Asp Ile Asp Val Glu Trp Gln Ser
Asn Gly Gln Gln Glu145 150 155
160Pro Glu Arg Lys His Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu
Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln 180
185 190Gln Gly Asp Pro Phe Thr Cys Ala Val Met His
Glu Thr Leu Gln Asn 195 200 205His
Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys 210
215 2202220PRTUnknownWild-type canine IgG-B Fc , Protein
A +, C1q +, CD16 + 2Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg
Glu 50 55 60Glu Gln Phe Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr
Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Gln Ala His Gln Pro Ser
Val Tyr Val Leu Pro Pro Ser Arg Glu Glu 115 120
125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp
Phe Phe 130 135 140Pro Pro Asp Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys
210 215 2203220PRTUnknownWild-type canine
IgG-C Fc , Protein A - , C1q +, CD16 + 3Pro Gly Cys Gly Leu Leu Gly
Gly Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr
Val Thr Cys 20 25 30Val Val
Val Asp Leu Asp Pro Glu Asn Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Ser Lys Gln Val Gln Thr Ala
Asn Thr Gln Pro Arg Glu 50 55 60Glu
Gln Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Ser
Gly Lys Gln Phe Lys Cys Lys Val Asn Asn 85
90 95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser
Lys Thr Pro Gly 100 105 110Gln
Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr
Cys Leu Val Lys Asp Phe Phe 130 135
140Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr
Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp
Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His
195 200 205Tyr Thr Gln Ile Ser Leu Ser
His Ser Pro Gly Lys 210 215
2204221PRTUnknownWild-type canine IgG-D Fc , Protein A -, C1q -,
CD16 4Pro Val Pro Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Ile
Leu Arg Ile Thr Arg Thr Pro Glu Ile Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val
Gln Ile Ser Trp 35 40 45Phe Val
Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Gln Gln Phe Asn Ser Thr Tyr Arg Val Val Ser
Val Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His
85 90 95Ile Gly Leu Pro Ser
Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro
Ser Pro Lys Glu 115 120 125Leu Ser
Ser Ser Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe 130
135 140Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser
Asn Gly Gln Pro Glu145 150 155
160Pro Glu Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu
Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln 180
185 190Gln Gly Asp Thr Phe Thr Cys Ala Val Met His
Glu Ala Leu Gln Asn 195 200 205His
Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys 210
215 2205221PRTArtificial SequenceSynthetic Variant canine
IgG-A Fc , C1q -, Protein A +, I(21)T, R(23)L, T(25)A, E(80)G,
T(205)A, Q(207)H 5Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val Leu Ile Phe
Pro Pro1 5 10 15Lys Pro
Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu Asp
Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg Glu 50
55 60Gln Gln Phe Asn Gly Thr Tyr Arg Val
Val Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val
Asn His 85 90 95Ile Asp
Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Arg Ala His Lys Pro Ser Val Tyr Val
Leu Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Ser Ile Thr Cys Leu Ile Lys Asp Phe
130 135 140Tyr Pro Pro Asp Ile Asp Val
Glu Trp Gln Ser Asn Gly Gln Gln Glu145 150
155 160Pro Glu Arg Lys His Arg Met Thr Pro Pro Gln Leu
Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Pro Phe Thr
Cys Ala Val Met His Glu Ala Leu His Asn 195 200
205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
210 215 2206220PRTArtificial
SequenceSynthetic Variant canine IgG-C Fc , C1q +, Protein A +,
I(21)T, V(23)L, T(24)I 6Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg
Glu 50 55 60Glu Gln Ser Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys
Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro Asn
Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp
Phe Phe 130 135 140Pro Pro Glu Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys
210 215 2207221PRTArtificial
SequenceSynthetic Variant canine IgG-D Fc , C1q -, Protein A +,
I(21)T, R(23)L, T(25)A, E(80)G, Q(207)H 7Pro Val Pro Glu Ser Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Ile
Thr Cys 20 25 30Val Val Leu
Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys
Thr Gln Pro Arg Glu 50 55 60Gln Gln
Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Thr Gly
Lys Glu Phe Lys Cys Arg Val Asn His 85 90
95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys
Ala Arg Gly 100 105 110Gln Ala
His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu 115
120 125Leu Ser Ser Ser Asp Thr Val Thr Leu Thr
Cys Leu Ile Lys Asp Phe 130 135 140Phe
Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro Glu145
150 155 160Pro Glu Ser Lys Tyr His
Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly 165
170 175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys
Ser Arg Trp Gln 180 185 190Gln
Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu His Asn 195
200 205His Tyr Thr Asp Leu Ser Leu Ser His
Ser Pro Gly Lys 210 215
2208221PRTArtificial SequenceSynthetic Variant canine IgG-A Fc ,
Heterodimer chain 1 T(138)Y 8Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val
Leu Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Val Thr Cys
20 25 30Val Val Leu Asp Leu Gly Arg
Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg
Glu 50 55 60Gln Gln Phe Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Glu65 70
75 80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys
Cys Arg Val Asn His 85 90
95Ile Asp Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Arg Ala His Lys Pro Ser
Val Tyr Val Leu Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Ser Ile Tyr Cys Leu Ile Lys
Asp Phe 130 135 140Tyr Pro Pro Asp Ile
Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu145 150
155 160Pro Glu Arg Lys His Arg Met Thr Pro Pro
Gln Leu Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Pro Phe
Thr Cys Ala Val Met His Glu Thr Leu Gln Asn 195
200 205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly
Lys 210 215 2209221PRTArtificial
SequenceSynthetic Variant canine IgG-A Fc , Heterodimer chain 2,
Y(181)T 9Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val Leu Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp
Ile Leu Arg Ile Thr Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu
Val Gln Ile Ser Trp 35 40 45Phe
Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg Glu 50
55 60Gln Gln Phe Asn Gly Thr Tyr Arg Val Val
Ser Val Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn
His 85 90 95Ile Asp Leu
Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Arg Ala His Lys Pro Ser Val Tyr Val Leu
Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Ser Ile Thr Cys Leu Ile Lys Asp Phe 130
135 140Tyr Pro Pro Asp Ile Asp Val Glu
Trp Gln Ser Asn Gly Gln Gln Glu145 150
155 160Pro Glu Arg Lys His Arg Met Thr Pro Pro Gln Leu
Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Thr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Pro Phe Thr
Cys Ala Val Met His Glu Thr Leu Gln Asn 195 200
205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
210 215 22010220PRTArtificial
SequenceSynthetic Variant canine IgG-B Fc , Heterodimer chain 1
T(137)Y 10Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro1 5 10 15Lys Pro Lys
Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu Asp Pro
Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Glu Gln Phe Asn Gly Thr Tyr Arg Val
Val Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Lys Val
Asn Asn 85 90 95Lys Ala
Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val
Leu Pro Pro Ser Arg Glu Glu 115 120
125Leu Ser Lys Asn Thr Val Ser Leu Tyr Cys Leu Ile Lys Asp Phe Phe
130 135 140Pro Pro Asp Ile Asp Val Glu
Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp
Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210
215 22011220PRTArtificial SequenceSynthetic
Variant canine IgG-B Fc , Heterodimer chain 2, Y(180)T 11Pro Ala Pro
Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Thr Leu Leu Ile Ala
Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Gln Met
Gln Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp
Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn 85
90 95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu
115 120 125Leu Ser Lys Asn Thr Val Ser
Leu Thr Cys Leu Ile Lys Asp Phe Phe 130 135
140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Thr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Glu
Ser Leu Ser His Ser Pro Gly Lys 210 215
22012220PRTArtificial SequenceSynthetic Variant canine IgG-C Fc ,
Heterodimer chain 1 T(137)Y 12Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val
Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg
Glu 50 55 60Glu Gln Ser Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys
Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro Asn
Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Tyr Cys Leu Val Lys Asp
Phe Phe 130 135 140Pro Pro Glu Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys
210 215 22013220PRTArtificial
SequenceSynthetic Variant canine IgG-C Fc , Heterodimer chain 2,
Y(180)T 13Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro1 5 10 15Lys Pro Lys
Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu Asn Pro
Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50
55 60Glu Gln Ser Asn Gly Thr Tyr Arg Val
Val Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val
Asn Asn 85 90 95Lys Ala
Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly 100
105 110Gln Ala His Gln Pro Asn Val Tyr Val
Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe
130 135 140Pro Pro Glu Ile Asp Val Glu
Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp
Glu Asp Gly Ser 165 170
175Tyr Phe Leu Thr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210
215 22014221PRTArtificial SequenceSynthetic
Variant canine IgG-D Fc , Heterodimer chain 1 T(138)Y 14Pro Val Pro
Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Arg Ile Thr
Arg Thr Pro Glu Ile Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Glu Val
His Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Gln Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65
70 75 80His Gln Asp Trp
Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu
115 120 125Leu Ser Ser Ser Asp Thr Val
Thr Leu Tyr Cys Leu Ile Lys Asp Phe 130 135
140Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro
Glu145 150 155 160Pro Glu
Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln 180 185
190Gln Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu
Gln Asn 195 200 205His Tyr Thr Asp
Leu Ser Leu Ser His Ser Pro Gly Lys 210 215
22015221PRTArtificial SequenceSynthetic Variant canine IgG-D Fc ,
Heterodimer chain 2, Y(181)T 15Pro Val Pro Glu Ser Leu Gly Gly Pro Ser
Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Ile Thr Cys
20 25 30Val Val Leu Asp Leu Gly
Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro
Arg Glu 50 55 60Gln Gln Phe Asn Ser
Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65 70
75 80His Gln Asp Trp Leu Thr Gly Lys Glu Phe
Lys Cys Arg Val Asn His 85 90
95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Gln Ala His Gln Pro
Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu 115
120 125Leu Ser Ser Ser Asp Thr Val Thr Leu Thr Cys Leu
Ile Lys Asp Phe 130 135 140Phe Pro Pro
Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro Glu145
150 155 160Pro Glu Ser Lys Tyr His Thr
Thr Ala Pro Gln Leu Asp Glu Asp Gly 165
170 175Ser Tyr Phe Leu Thr Ser Lys Leu Ser Val Asp Lys
Ser Arg Trp Gln 180 185 190Gln
Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu Gln Asn 195
200 205His Tyr Thr Asp Leu Ser Leu Ser His
Ser Pro Gly Lys 210 215
22016237PRTUnknownWild-type feline IgG2 Fc, Protein A +, C1q - 16Pro Lys
Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Lys1 5
10 15Cys Pro Val Pro Glu Ile Pro Gly
Ala Pro Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val
Thr 35 40 45Cys Leu Val Val Asp
Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg
Pro Arg65 70 75 80Glu
Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu Lys
Gly Lys Glu Phe Lys Cys Lys Val Asn 100 105
110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys
Ala Lys 115 120 125Gly Gln Pro His
Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu
Ile Lys Gly Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn Tyr
Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg
Ser His Trp Gln 195 200 205Arg Gly
Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro
Gly Lys225 230 23517237PRTArtificial
SequenceSynthetic Variant feline IgG2 Fc , Hinge Cys, G(14)C 17Pro
Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Cys Pro Lys1
5 10 15Cys Pro Val Pro Glu Ile Pro
Gly Ala Pro Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr
Arg Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Gly Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 23518228PRTArtificial
SequenceSynthetic Variant equine Fc IgG2 (with equine IgG1 hinge),
Protein A - , C1q - 18Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu
Leu Gly Gly1 5 10 15Pro
Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala Leu Met Ile 20
25 30Ser Arg Thr Pro Val Val Thr Cys
Val Val Val Asn Leu Ser Asp Gln 35 40
45Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val His
50 55 60Ser Ala Ile Thr Lys Gln Arg Glu
Ala Gln Phe Asn Ser Thr Tyr Arg65 70 75
80Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu
Ser Gly Lys 85 90 95Glu
Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile Ser
100 105 110Arg Ala Ile Ser Arg Gly Lys
Gly Pro Ser Arg Val Pro Gln Val Tyr 115 120
125Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
Val 130 135 140Thr Cys Leu Val Lys Asp
Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp145 150
155 160Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys
Tyr Ser Thr Thr Pro 165 170
175Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu Ser
180 185 190Leu Glu Thr Ser Arg Trp
Gln Gln Val Glu Ser Phe Thr Cys Ala Val 195 200
205Met His Glu Ala Leu His Asn His Phe Thr Lys Thr Asp Ile
Ser Glu 210 215 220Ser Leu Gly
Lys22519228PRTArtificial SequenceSynthetic Variant equine IgG2 Fc (with
equine IgG1 hinge), C1q -, Protein A +, A(29)T, F(217)Y 19Asp Met
Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu Leu Gly Gly1 5
10 15Pro Ser Val Phe Ile Phe Pro Pro
Asn Pro Lys Asp Thr Leu Met Ile 20 25
30Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser Asp
Gln 35 40 45Tyr Pro Asp Val Gln
Phe Ser Trp Tyr Val Asp Asn Thr Glu Val His 50 55
60Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr
Tyr Arg65 70 75 80Val
Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser Gly Lys
85 90 95Glu Phe Lys Cys Ser Val Thr
Asn Val Gly Val Pro Gln Pro Ile Ser 100 105
110Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln
Val Tyr 115 120 125Val Leu Pro Pro
His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser Val 130
135 140Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile
Ser Val Glu Trp145 150 155
160Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr Thr Pro
165 170 175Ala Gln Leu Asp Gly
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu Ser 180
185 190Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe
Thr Cys Ala Val 195 200 205Met His
Glu Ala Leu His Asn His Tyr Thr Lys Thr Asp Ile Ser Glu 210
215 220Ser Leu Gly Lys2252029PRTArtificial
SequenceSynthetic Variant GLP1 (7-35)misc_feature(2)..(2)X may be G or S
20His Xaa Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1
5 10 15Gln Ala Ala Lys Glu Phe
Ile Ala Trp Leu Val Lys Gly 20
252129PRTArtificial SequenceSynthetic Glucagon (Gluc) 21His Ser Gln Gly
Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser1 5
10 15Arg Arg Ala Gln Asp Phe Val Gln Trp Leu
Met Asn Thr 20 252239PRTArtificial
SequenceSynthetic Extendin-4 22His Gly Glu Gly Thr Phe Thr Ser Asp Leu
Ser Lys Gln Met Glu Glu1 5 10
15Glu Ala Val Arg Leu Phe Ile Glu Trp Leu Lys Asn Gly Gly Pro Ser
20 25 30Ser Gly Ala Pro Pro Pro
Ser 3523325PRTArtificial SequenceSynthetic GLP1-G8/GLP-2G_III_
WTfeIgG2 23His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu
Gly1 5 10 15Gln Ala Ala
Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly Gly 20
25 30Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
Ser Gly Gly Gly Gly Ser 35 40
45Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Lys 50
55 60Cys Pro Val Pro Glu Ile Pro Gly Ala
Pro Ser Val Phe Ile Phe Pro65 70 75
80Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 85 90 95Cys Leu
Val Val Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 100
105 110Trp Phe Val Asp Asn Thr Glu Met His
Thr Ala Lys Thr Arg Pro Arg 115 120
125Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
130 135 140Leu His Gln Asp Trp Leu Lys
Gly Lys Glu Phe Lys Cys Lys Val Asn145 150
155 160Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile
Ser Lys Ala Lys 165 170
175Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu
180 185 190Glu Leu Ser Glu Asn Lys
Val Ser Val Thr Cys Leu Ile Lys Gly Phe 195 200
205His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln
Pro Glu 210 215 220Pro Glu Asn Asn Tyr
Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly225 230
235 240Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val
Asp Arg Ser His Trp Gln 245 250
255Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser
260 265 270His His Thr Gln Lys
Ser Leu Thr Gln Ser Pro Gly Lys Gly Gly Gly 275
280 285Gly Ser Gly Gly Gly Gly His Ala Glu Gly Thr Phe
Thr Ser Asp Val 290 295 300Ser Ser Tyr
Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu305
310 315 320Val Lys Gly Gly Gly
32524285PRTArtificial SequenceSynthetic GLP1-G8_I_WTfeIgG2 24His Gly
Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala
Trp Leu Val Lys Gly Gly Gly Gly 20 25
30Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
Ser 35 40 45Pro Lys Thr Ala Ser
Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Lys 50 55
60Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe Ile
Phe Pro65 70 75 80Pro
Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
85 90 95Cys Leu Val Val Asp Leu Gly
Pro Asp Asp Ser Asn Val Gln Ile Thr 100 105
110Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg
Pro Arg 115 120 125Glu Glu Gln Phe
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile 130
135 140Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys
Cys Lys Val Asn145 150 155
160Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys Ala Lys
165 170 175Gly Gln Pro His Glu
Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 180
185 190Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu
Ile Lys Gly Phe 195 200 205His Pro
Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu 210
215 220Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln
Leu Asp Ser Asp Gly225 230 235
240Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp Gln
245 250 255Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 260
265 270His His Thr Gln Lys Ser Leu Thr Gln Ser Pro
Gly Lys 275 280
28525325PRTArtificial SequenceSynthetic GLP1-G8/GLP1-2G_III_ VARfeIgG2
25His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1
5 10 15Gln Ala Ala Lys Glu Phe
Ile Ala Trp Leu Val Lys Gly Gly Gly Gly 20 25
30Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly
Gly Gly Ser 35 40 45Pro Lys Thr
Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Cys Pro Lys 50
55 60Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val
Phe Ile Phe Pro65 70 75
80Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
85 90 95Cys Leu Val Val Asp Leu
Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 100
105 110Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys
Thr Arg Pro Arg 115 120 125Glu Glu
Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile 130
135 140Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe
Lys Cys Lys Val Asn145 150 155
160Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys Ala Lys
165 170 175Gly Gln Pro His
Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 180
185 190Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Gly Phe 195 200 205His
Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu 210
215 220Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro
Gln Leu Asp Ser Asp Gly225 230 235
240Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp
Gln 245 250 255Arg Gly Asn
Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 260
265 270His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys Gly Gly Gly 275 280
285Gly Ser Gly Gly Gly Gly His Ala Glu Gly Thr Phe Thr Ser Asp Val 290
295 300Ser Ser Tyr Leu Glu Gly Gln Ala
Ala Lys Glu Phe Ile Ala Trp Leu305 310
315 320Val Lys Gly Gly Gly
32526285PRTArtificial SequenceSynthetic GLP1-G8_I_VARfeIgG2 26His Gly Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp
Leu Val Lys Gly Gly Gly Gly 20 25
30Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
35 40 45Pro Lys Thr Ala Ser Thr Ile
Glu Ser Lys Thr Gly Glu Cys Pro Lys 50 55
60Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe Ile Phe Pro65
70 75 80Pro Lys Pro Lys
Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 85
90 95Cys Leu Val Val Asp Leu Gly Pro Asp Asp
Ser Asn Val Gln Ile Thr 100 105
110Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg Pro Arg
115 120 125Glu Glu Gln Phe Asn Ser Thr
Tyr Arg Val Val Ser Val Leu Pro Ile 130 135
140Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val
Asn145 150 155 160Ser Lys
Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys Ala Lys
165 170 175Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro Pro Thr Gln Glu 180 185
190Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys
Gly Phe 195 200 205His Pro Pro Asp
Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu 210
215 220Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly225 230 235
240Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp Gln
245 250 255Arg Gly Asn Thr Tyr
Thr Cys Ser Val Ser His Glu Ala Leu His Ser 260
265 270His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly
Lys 275 280 28527327PRTArtificial
SequenceSynthetic GLP1-S8/GLP1-3G_III_ WTfeIgG2 27His Ser Glu Gly Thr Phe
Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys
Gly Gly Ala Gly 20 25 30Gly
Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35
40 45Ser Pro Lys Thr Ala Ser Thr Ile Glu
Ser Lys Thr Gly Glu Gly Pro 50 55
60Lys Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe Ile Phe65
70 75 80Pro Pro Lys Pro Lys
Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val 85
90 95Thr Cys Leu Val Val Asp Leu Gly Pro Asp Asp
Ser Asn Val Gln Ile 100 105
110Thr Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg Pro
115 120 125Arg Glu Glu Gln Phe Asn Ser
Thr Tyr Arg Val Val Ser Val Leu Pro 130 135
140Ile Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys
Val145 150 155 160Asn Ser
Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys Ala
165 170 175Lys Gly Gln Pro His Glu Pro
Gln Val Tyr Val Leu Pro Pro Thr Gln 180 185
190Glu Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile
Lys Gly 195 200 205Phe His Pro Pro
Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro 210
215 220Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln
Leu Asp Ser Asp225 230 235
240Gly Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp
245 250 255Gln Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His 260
265 270Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro
Gly Lys Gly Gly 275 280 285Gly Gly
Ser Gly Gly Gly Gly His Ala Glu Gly Thr Phe Thr Ser Asp 290
295 300Val Ser Ser Tyr Leu Glu Gly Gln Ala Ala Lys
Glu Phe Ile Ala Trp305 310 315
320Leu Val Lys Gly Gly Gly Gly 32528286PRTArtificial
SequenceSynthetic GLP1-S8_I_WTfeIgG2 28His Ser Glu Gly Thr Phe Thr Ser
Asp Val Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly
Ala Gly 20 25 30Gly Gly Gly
Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35
40 45Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys
Thr Gly Glu Gly Pro 50 55 60Lys Cys
Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe Ile Phe65
70 75 80Pro Pro Lys Pro Lys Asp Thr
Leu Ser Ile Ser Arg Thr Pro Glu Val 85 90
95Thr Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn
Val Gln Ile 100 105 110Thr Trp
Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg Pro 115
120 125Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg
Val Val Ser Val Leu Pro 130 135 140Ile
Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val145
150 155 160Asn Ser Lys Ser Leu Pro
Ser Ala Met Glu Arg Thr Ile Ser Lys Ala 165
170 175Lys Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu
Pro Pro Thr Gln 180 185 190Glu
Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Gly 195
200 205Phe His Pro Pro Asp Ile Ala Val Glu
Trp Glu Ile Thr Gly Gln Pro 210 215
220Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp225
230 235 240Gly Thr Tyr Phe
Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp 245
250 255Gln Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His 260 265
270Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys 275
280 28529323PRTArtificial SequenceSynthetic
GLP1-G8/GLP1-3G_III_ VARcaIgGD 29His Gly Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Pro Lys Glu Ser Thr Ser Lys Cys Ile Ser Pro Cys Pro
Val Pro 50 55 60Glu Ser Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys65 70
75 80Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu
Ile Thr Cys Val Val Leu 85 90
95Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp Phe Val Asp
100 105 110Gly Lys Glu Val His
Thr Ala Lys Thr Gln Pro Arg Glu Gln Gln Phe 115
120 125Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
Gly His Gln Asp 130 135 140Trp Leu Thr
Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile Gly Leu145
150 155 160Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly Gln Ala His 165
170 175Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys
Glu Leu Ser Ser 180 185 190Ser
Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe Pro Pro 195
200 205Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Pro Glu Pro Glu Ser 210 215
220Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly Ser Tyr Phe225
230 235 240Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly Asp 245
250 255Thr Phe Thr Cys Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr 260 265
270Asp Leu Ser Leu Ser His Ser Pro Gly Lys Gly Gly Gly Gly Ser Gly
275 280 285Gly Gly Gly His Ala Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr 290 295
300Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys
Gly305 310 315 320Gly Gly
Gly30282PRTArtificial SequenceSynthetic GLP1-G8_I_VARcaIgGD 30His Gly Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp
Leu Val Lys Gly Gly Ala Gly 20 25
30Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
35 40 45Ser Pro Lys Glu Ser Thr Cys
Lys Cys Ile Ser Pro Cys Pro Val Pro 50 55
60Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys65
70 75 80Asp Thr Leu Leu
Ile Ala Arg Thr Pro Glu Ile Thr Cys Val Val Leu 85
90 95Asp Leu Gly Arg Glu Asp Pro Glu Val Gln
Ile Ser Trp Phe Val Asp 100 105
110Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu Gln Gln Phe
115 120 125Asn Ser Thr Tyr Arg Val Val
Ser Val Leu Pro Ile Gly His Gln Asp 130 135
140Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile Gly
Leu145 150 155 160Pro Ser
Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly Gln Ala His
165 170 175Gln Pro Ser Val Tyr Val Leu
Pro Pro Ser Pro Lys Glu Leu Ser Ser 180 185
190Ser Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe
Pro Pro 195 200 205Glu Ile Asp Val
Glu Trp Gln Ser Asn Gly Gln Pro Glu Pro Glu Ser 210
215 220Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp
Gly Ser Tyr Phe225 230 235
240Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly Asp
245 250 255Thr Phe Thr Cys Ala
Val Met His Glu Ala Leu His Asn His Tyr Thr 260
265 270Asp Leu Ser Leu Ser His Ser Pro Gly Lys
275 28031323PRTArtificial SequenceSynthetic
GLP1-S8/GLP1-3G_III_ VARcaIgGD 31His Ser Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Pro Lys Glu Ser Thr Ser Lys Cys Ile Ser Pro Cys Pro
Val Pro 50 55 60Glu Ser Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys65 70
75 80Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu
Ile Thr Cys Val Val Leu 85 90
95Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp Phe Val Asp
100 105 110Gly Lys Glu Val His
Thr Ala Lys Thr Gln Pro Arg Glu Gln Gln Phe 115
120 125Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
Gly His Gln Asp 130 135 140Trp Leu Thr
Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile Gly Leu145
150 155 160Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly Gln Ala His 165
170 175Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys
Glu Leu Ser Ser 180 185 190Ser
Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe Pro Pro 195
200 205Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Pro Glu Pro Glu Ser 210 215
220Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly Ser Tyr Phe225
230 235 240Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly Asp 245
250 255Thr Phe Thr Cys Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr 260 265
270Asp Leu Ser Leu Ser His Ser Pro Gly Lys Gly Gly Gly Gly Ser Gly
275 280 285Gly Gly Gly His Ala Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr 290 295
300Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys
Gly305 310 315 320Gly Gly
Gly32282PRTArtificial SequenceSynthetic GLP1-S8_I_VARcaIgGD 32His Ser Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp
Leu Val Lys Gly Gly Ala Gly 20 25
30Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
35 40 45Ser Pro Lys Glu Ser Thr Cys
Lys Cys Ile Ser Pro Cys Pro Val Pro 50 55
60Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys65
70 75 80Asp Thr Leu Leu
Ile Ala Arg Thr Pro Glu Ile Thr Cys Val Val Leu 85
90 95Asp Leu Gly Arg Glu Asp Pro Glu Val Gln
Ile Ser Trp Phe Val Asp 100 105
110Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu Gln Gln Phe
115 120 125Asn Ser Thr Tyr Arg Val Val
Ser Val Leu Pro Ile Gly His Gln Asp 130 135
140Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile Gly
Leu145 150 155 160Pro Ser
Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly Gln Ala His
165 170 175Gln Pro Ser Val Tyr Val Leu
Pro Pro Ser Pro Lys Glu Leu Ser Ser 180 185
190Ser Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe
Pro Pro 195 200 205Glu Ile Asp Val
Glu Trp Gln Ser Asn Gly Gln Pro Glu Pro Glu Ser 210
215 220Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp
Gly Ser Tyr Phe225 230 235
240Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly Asp
245 250 255Thr Phe Thr Cys Ala
Val Met His Glu Ala Leu His Asn His Tyr Thr 260
265 270Asp Leu Ser Leu Ser His Ser Pro Gly Lys
275 28033318PRTArtificial SequenceSynthetic
GLP1-G8/GLPL1-3G_III_ VAReqIgG2 33His Gly Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu
Leu Gly 50 55 60Gly Pro Ser Val Phe
Ile Phe Pro Pro Asn Pro Lys Asp Thr Leu Met65 70
75 80Ile Ser Arg Thr Pro Val Val Thr Cys Val
Val Val Asn Leu Ser Asp 85 90
95Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val
100 105 110His Ser Ala Ile Thr
Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr 115
120 125Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp
Trp Leu Ser Gly 130 135 140Lys Glu Phe
Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile145
150 155 160Ser Arg Ala Ile Ser Arg Gly
Lys Gly Pro Ser Arg Val Pro Gln Val 165
170 175Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys
Ser Lys Val Ser 180 185 190Val
Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu 195
200 205Trp Gln Ser Asn Arg Trp Pro Glu Leu
Glu Gly Lys Tyr Ser Thr Thr 210 215
220Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu225
230 235 240Ser Leu Glu Thr
Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala 245
250 255Val Met His Glu Ala Leu His Asn His Tyr
Thr Lys Thr Asp Ile Ser 260 265
270Glu Ser Leu Gly Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly His Ala
275 280 285Glu Gly Thr Phe Thr Ser Asp
Val Ser Ser Tyr Leu Glu Gly Gln Ala 290 295
300Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly Gly305
310 31534277PRTArtificial SequenceSynthetic
GLP1-G8_I_VAReqIgG2 34His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr
Leu Glu Gly1 5 10 15Gln
Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly 20
25 30Gly Gly Gly Gly Ser Gly Gly Gly
Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu Leu Gly
50 55 60Gly Pro Ser Val Phe Ile Phe Pro
Pro Asn Pro Lys Asp Thr Leu Met65 70 75
80Ile Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn
Leu Ser Asp 85 90 95Gln
Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val
100 105 110His Ser Ala Ile Thr Lys Gln
Arg Glu Ala Gln Phe Asn Ser Thr Tyr 115 120
125Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser
Gly 130 135 140Lys Glu Phe Lys Cys Ser
Val Thr Asn Val Gly Val Pro Gln Pro Ile145 150
155 160Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser
Arg Val Pro Gln Val 165 170
175Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
180 185 190Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu 195 200
205Trp Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser
Thr Thr 210 215 220Pro Ala Gln Leu Asp
Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu225 230
235 240Ser Leu Glu Thr Ser Arg Trp Gln Gln Val
Glu Ser Phe Thr Cys Ala 245 250
255Val Met His Glu Ala Leu His Asn His Tyr Thr Lys Thr Asp Ile Ser
260 265 270Glu Ser Leu Gly Lys
27535318PRTArtificial SequenceSynthetic GLP1-S8/GLP1-3G_III_
VAReqIgG2 35His Ser Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu
Gly1 5 10 15Gln Ala Ala
Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly 20
25 30Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly
Gly Gly Ser Gly Gly Gly 35 40
45Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu Leu Gly 50
55 60Gly Pro Ser Val Phe Ile Phe Pro Pro
Asn Pro Lys Asp Thr Leu Met65 70 75
80Ile Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu
Ser Asp 85 90 95Gln Tyr
Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val 100
105 110His Ser Ala Ile Thr Lys Gln Arg Glu
Ala Gln Phe Asn Ser Thr Tyr 115 120
125Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser Gly
130 135 140Lys Glu Phe Lys Cys Ser Val
Thr Asn Val Gly Val Pro Gln Pro Ile145 150
155 160Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg
Val Pro Gln Val 165 170
175Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
180 185 190Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu 195 200
205Trp Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser
Thr Thr 210 215 220Pro Ala Gln Leu Asp
Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu225 230
235 240Ser Leu Glu Thr Ser Arg Trp Gln Gln Val
Glu Ser Phe Thr Cys Ala 245 250
255Val Met His Glu Ala Leu His Asn His Tyr Thr Lys Thr Asp Ile Ser
260 265 270Glu Ser Leu Gly Lys
Gly Gly Gly Gly Ser Gly Gly Gly Gly His Ala 275
280 285Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu
Glu Gly Gln Ala 290 295 300Ala Lys Glu
Phe Ile Ala Trp Leu Val Lys Gly Gly Gly Gly305 310
31536277PRTArtificial SequenceSynthetic GLP1-S8_I_VAReqIgG2
36His Ser Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1
5 10 15Gln Ala Ala Lys Glu Phe
Ile Ala Trp Leu Val Lys Gly Gly Ala Gly 20 25
30Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser
Gly Gly Gly 35 40 45Ser Asp Met
Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu Leu Gly 50
55 60Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys
Asp Thr Leu Met65 70 75
80Ile Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser Asp
85 90 95Gln Tyr Pro Asp Val Gln
Phe Ser Trp Tyr Val Asp Asn Thr Glu Val 100
105 110His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe
Asn Ser Thr Tyr 115 120 125Arg Val
Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser Gly 130
135 140Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly
Val Pro Gln Pro Ile145 150 155
160Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln Val
165 170 175Tyr Val Leu Pro
Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser 180
185 190Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Asp Ile Ser Val Glu 195 200 205Trp
Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr Thr 210
215 220Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr
Phe Leu Tyr Ser Lys Leu225 230 235
240Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys
Ala 245 250 255Val Met His
Glu Ala Leu His Asn His Tyr Thr Lys Thr Asp Ile Ser 260
265 270Glu Ser Leu Gly Lys
27537346PRTArtificial SequenceSynthetic ssGLP1-S8/GLP1-3G_III_ WTfeIgG2
37Met Ala Val Leu Gly Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1
5 10 15Val Leu Ser His Ser Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu
Val Lys Gly 35 40 45Gly Ala Gly
Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser 50
55 60Gly Gly Gly Ser Pro Lys Thr Ala Ser Thr Ile Glu
Ser Lys Thr Gly65 70 75
80Glu Gly Pro Lys Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val
85 90 95Phe Ile Phe Pro Pro Lys
Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr 100
105 110Pro Glu Val Thr Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asn 115 120 125Val Gln
Ile Thr Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys 130
135 140Thr Arg Pro Arg Glu Glu Gln Phe Asn Ser Thr
Tyr Arg Val Val Ser145 150 155
160Val Leu Pro Ile Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys
165 170 175Cys Lys Val Asn
Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile 180
185 190Ser Lys Ala Lys Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro 195 200 205Pro
Thr Gln Glu Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu 210
215 220Ile Lys Gly Phe His Pro Pro Asp Ile Ala
Val Glu Trp Glu Ile Thr225 230 235
240Gly Gln Pro Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln
Leu 245 250 255Asp Ser Asp
Gly Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg 260
265 270Ser His Trp Gln Arg Gly Asn Thr Tyr Thr
Cys Ser Val Ser His Glu 275 280
285Ala Leu His Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly 290
295 300Lys Gly Gly Gly Gly Ser Gly Gly
Gly Gly His Ala Glu Gly Thr Phe305 310
315 320Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly Gln Ala
Ala Lys Glu Phe 325 330
335Ile Ala Trp Leu Val Lys Gly Gly Gly Gly 340
34538344PRTArtificial SequenceSynthetic ssGLP1-G8/GLP1-2G_III_ VARfeIgG2
38Met Ala Val Leu Gly Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1
5 10 15Val Leu Ser His Gly Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu
Val Lys Gly 35 40 45Gly Gly Gly
Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly 50
55 60Gly Gly Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser
Lys Thr Gly Glu65 70 75
80Cys Pro Lys Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe
85 90 95Ile Phe Pro Pro Lys Pro
Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro 100
105 110Glu Val Thr Cys Leu Val Val Asp Leu Gly Pro Asp
Asp Ser Asn Val 115 120 125Gln Ile
Thr Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr 130
135 140Arg Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr
Arg Val Val Ser Val145 150 155
160Leu Pro Ile Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys
165 170 175Lys Val Asn Ser
Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser 180
185 190Lys Ala Lys Gly Gln Pro His Glu Pro Gln Val
Tyr Val Leu Pro Pro 195 200 205Thr
Gln Glu Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile 210
215 220Lys Gly Phe His Pro Pro Asp Ile Ala Val
Glu Trp Glu Ile Thr Gly225 230 235
240Gln Pro Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu
Asp 245 250 255Ser Asp Gly
Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser 260
265 270His Trp Gln Arg Gly Asn Thr Tyr Thr Cys
Ser Val Ser His Glu Ala 275 280
285Leu His Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys 290
295 300Gly Gly Gly Gly Ser Gly Gly Gly
Gly His Ala Glu Gly Thr Phe Thr305 310
315 320Ser Asp Val Ser Ser Tyr Leu Glu Gly Gln Ala Ala
Lys Glu Phe Ile 325 330
335Ala Trp Leu Val Lys Gly Gly Gly 34039304PRTArtificial
SequenceSynthetic ssGLP1-G8_I_VARfeIgG2 39Met Ala Val Leu Gly Leu Leu Phe
Cys Leu Val Thr Phe Pro Ser Cys1 5 10
15Val Leu Ser His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser
Ser Tyr 20 25 30Leu Glu Gly
Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly 35
40 45Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly
Gly Gly Ser Gly Gly 50 55 60Gly Gly
Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu65
70 75 80Cys Pro Lys Cys Pro Val Pro
Glu Ile Pro Gly Ala Pro Ser Val Phe 85 90
95Ile Phe Pro Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser
Arg Thr Pro 100 105 110Glu Val
Thr Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn Val 115
120 125Gln Ile Thr Trp Phe Val Asp Asn Thr Glu
Met His Thr Ala Lys Thr 130 135 140Arg
Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val145
150 155 160Leu Pro Ile Leu His Gln
Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys 165
170 175Lys Val Asn Ser Lys Ser Leu Pro Ser Ala Met Glu
Arg Thr Ile Ser 180 185 190Lys
Ala Lys Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro 195
200 205Thr Gln Glu Glu Leu Ser Glu Asn Lys
Val Ser Val Thr Cys Leu Ile 210 215
220Lys Gly Phe His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly225
230 235 240Gln Pro Glu Pro
Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu Asp 245
250 255Ser Asp Gly Thr Tyr Phe Leu Tyr Ser Arg
Leu Ser Val Asp Arg Ser 260 265
270His Trp Gln Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala
275 280 285Leu His Ser His His Thr Gln
Lys Ser Leu Thr Gln Ser Pro Gly Lys 290 295
30040305PRTArtificial SequenceSynthetic ssGLP1-S8_I_WTfeIgG2 40Met
Ala Val Leu Gly Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1
5 10 15Val Leu Ser His Ser Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val
Lys Gly 35 40 45Gly Ala Gly Gly
Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser 50 55
60Gly Gly Gly Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser
Lys Thr Gly65 70 75
80Glu Gly Pro Lys Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val
85 90 95Phe Ile Phe Pro Pro Lys
Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr 100
105 110Pro Glu Val Thr Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asn 115 120 125Val Gln
Ile Thr Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys 130
135 140Thr Arg Pro Arg Glu Glu Gln Phe Asn Ser Thr
Tyr Arg Val Val Ser145 150 155
160Val Leu Pro Ile Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys
165 170 175Cys Lys Val Asn
Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile 180
185 190Ser Lys Ala Lys Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro 195 200 205Pro
Thr Gln Glu Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu 210
215 220Ile Lys Gly Phe His Pro Pro Asp Ile Ala
Val Glu Trp Glu Ile Thr225 230 235
240Gly Gln Pro Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln
Leu 245 250 255Asp Ser Asp
Gly Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg 260
265 270Ser His Trp Gln Arg Gly Asn Thr Tyr Thr
Cys Ser Val Ser His Glu 275 280
285Ala Leu His Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly 290
295 300Lys30541340PRTArtificial
SequenceSynthetic ssGLP1-G8/GLP1-2G_III_ VARcaIgGD 41Met Ala Val Leu Gly
Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1 5
10 15Val Leu Ser His Gly Glu Gly Thr Phe Thr Ser
Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly
35 40 45Gly Gly Gly Ser Gly Gly Gly Gly
Ser Gly Gly Gly Gly Ser Gly Gly 50 55
60Gly Gly Ser Pro Lys Glu Ser Thr Cys Lys Cys Ile Ser Pro Cys Pro65
70 75 80Val Pro Glu Ser Leu
Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys 85
90 95Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro
Glu Ile Thr Cys Val 100 105
110Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp Phe
115 120 125Val Asp Gly Lys Glu Val His
Thr Ala Lys Thr Gln Pro Arg Glu Gln 130 135
140Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly
His145 150 155 160Gln Asp
Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile
165 170 175Gly Leu Pro Ser Pro Ile Glu
Arg Thr Ile Ser Lys Ala Arg Gly Gln 180 185
190Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys
Glu Leu 195 200 205Ser Ser Ser Asp
Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe 210
215 220Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly
Gln Pro Glu Pro225 230 235
240Glu Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly Ser
245 250 255Tyr Phe Leu Tyr Ser
Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Gln 260
265 270Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala
Leu His Asn His 275 280 285Tyr Thr
Asp Leu Ser Leu Ser His Ser Pro Gly Lys Gly Gly Gly Gly 290
295 300Ser Gly Gly Gly Gly His Ala Glu Gly Thr Phe
Thr Ser Asp Val Ser305 310 315
320Ser Tyr Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val
325 330 335Lys Gly Gly Gly
34042340PRTArtificial SequenceSynthetic ssGLP1-G8/GLP1-2G_III_
VARcaIgGD 42Met Ala Val Leu Gly Leu Leu Phe Cys Leu Val Thr Phe Pro Ser
Cys1 5 10 15Val Leu Ser
His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr 20
25 30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile
Ala Trp Leu Val Lys Gly 35 40
45Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly 50
55 60Gly Gly Ser Pro Lys Glu Ser Thr Ser
Lys Cys Ile Ser Pro Cys Pro65 70 75
80Val Pro Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro Lys 85 90 95Pro Lys
Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Ile Thr Cys Val 100
105 110Val Leu Asp Leu Gly Arg Glu Asp Pro
Glu Val Gln Ile Ser Trp Phe 115 120
125Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu Gln
130 135 140Gln Phe Asn Ser Thr Tyr Arg
Val Val Ser Val Leu Pro Ile Gly His145 150
155 160Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg
Val Asn His Ile 165 170
175Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly Gln
180 185 190Ala His Gln Pro Ser Val
Tyr Val Leu Pro Pro Ser Pro Lys Glu Leu 195 200
205Ser Ser Ser Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp
Phe Phe 210 215 220Pro Pro Glu Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Pro Glu Pro225 230
235 240Glu Ser Lys Tyr His Thr Thr Ala Pro Gln
Leu Asp Glu Asp Gly Ser 245 250
255Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Gln
260 265 270Gly Asp Thr Phe Thr
Cys Ala Val Met His Glu Ala Leu His Asn His 275
280 285Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
Gly Gly Gly Gly 290 295 300Ser Gly Gly
Gly Gly His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser305
310 315 320Ser Tyr Leu Glu Gly Gln Ala
Ala Lys Glu Phe Ile Ala Trp Leu Val 325
330 335Lys Gly Gly Gly 34043342PRTArtificial
SequenceSynthetic ssGLP1-S8/GLP1-3G_III_ VARcaIgGD 43Met Ala Val Leu Gly
Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1 5
10 15Val Leu Ser His Ser Glu Gly Thr Phe Thr Ser
Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly
35 40 45Gly Ala Gly Gly Gly Gly Gly Ser
Gly Gly Gly Ser Gly Gly Gly Ser 50 55
60Gly Gly Gly Ser Pro Lys Glu Ser Thr Ser Lys Cys Ile Ser Pro Cys65
70 75 80Pro Val Pro Glu Ser
Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro 85
90 95Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr
Pro Glu Ile Thr Cys 100 105
110Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp
115 120 125Phe Val Asp Gly Lys Glu Val
His Thr Ala Lys Thr Gln Pro Arg Glu 130 135
140Gln Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
Gly145 150 155 160His Gln
Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His
165 170 175Ile Gly Leu Pro Ser Pro Ile
Glu Arg Thr Ile Ser Lys Ala Arg Gly 180 185
190Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro
Lys Glu 195 200 205Leu Ser Ser Ser
Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe 210
215 220Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Pro Glu225 230 235
240Pro Glu Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
245 250 255Ser Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln 260
265 270Gln Gly Asp Thr Phe Thr Cys Ala Val Met His Glu
Ala Leu His Asn 275 280 285His Tyr
Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys Gly Gly Gly 290
295 300Gly Ser Gly Gly Gly Gly His Ala Glu Gly Thr
Phe Thr Ser Asp Val305 310 315
320Ser Ser Tyr Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu
325 330 335Val Lys Gly Gly
Gly Gly 34044337PRTArtificial SequenceSynthetic
ssGLP1-G8/GLP1-3G_III_ VAReqIgG2 44Met Ala Val Leu Gly Leu Leu Phe Cys
Leu Val Thr Phe Pro Ser Cys1 5 10
15Val Leu Ser His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser Ser
Tyr 20 25 30Leu Glu Gly Gln
Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly 35
40 45Gly Ala Gly Gly Gly Gly Gly Ser Gly Gly Gly Ser
Gly Gly Gly Ser 50 55 60Gly Gly Gly
Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu65 70
75 80Leu Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro Asn Pro Lys Asp 85 90
95Thr Leu Met Ile Ser Arg Thr Pro Val Val Thr Cys Val Val
Val Asn 100 105 110Leu Ser Asp
Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn 115
120 125Thr Glu Val His Ser Ala Ile Thr Lys Gln Arg
Glu Ala Gln Phe Asn 130 135 140Ser Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp145
150 155 160Leu Ser Gly Lys Glu Phe Lys
Cys Ser Val Thr Asn Val Gly Val Pro 165
170 175Gln Pro Ile Ser Arg Ala Ile Ser Arg Gly Lys Gly
Pro Ser Arg Val 180 185 190Pro
Gln Val Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser 195
200 205Lys Val Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile 210 215
220Ser Val Glu Trp Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr225
230 235 240Ser Thr Thr Pro
Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr 245
250 255Ser Lys Leu Ser Leu Glu Thr Ser Arg Trp
Gln Gln Val Glu Ser Phe 260 265
270Thr Cys Ala Val Met His Glu Ala Leu His Asn His Tyr Thr Lys Thr
275 280 285Asp Ile Ser Glu Ser Leu Gly
Lys Gly Gly Gly Gly Ser Gly Gly Gly 290 295
300Gly His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu
Glu305 310 315 320Gly Gln
Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly
325 330 335Gly45298PRTArtificial
SequenceSynthetic ssGLP1-G8_I_VAReqIgG2 45Met Ala Val Leu Gly Leu Leu Phe
Cys Leu Val Thr Phe Pro Ser Cys1 5 10
15Val Leu Ser His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser
Ser Tyr 20 25 30Leu Glu Gly
Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly 35
40 45Gly Ala Gly Gly Gly Gly Gly Ser Gly Gly Gly
Ser Gly Gly Gly Ser 50 55 60Gly Gly
Gly Ser Ser Val Pro Lys Pro Gln Cys Pro Pro Tyr Thr His65
70 75 80Ser Lys Phe Leu Gly Gly Pro
Ser Val Phe Ile Phe Pro Pro Asn Pro 85 90
95Lys Asp Thr Leu Met Ile Ser Arg Thr Pro Val Val Thr
Cys Val Val 100 105 110Val Asn
Leu Ser Asp Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val 115
120 125Asp Asn Thr Glu Val His Ser Ala Ile Thr
Lys Gln Arg Glu Ala Gln 130 135 140Phe
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln145
150 155 160Asp Trp Leu Ser Gly Lys
Glu Phe Lys Cys Ser Val Thr Asn Val Gly 165
170 175Val Pro Gln Pro Ile Ser Arg Ala Ile Ser Arg Gly
Lys Gly Pro Ser 180 185 190Arg
Val Pro Gln Val Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala 195
200 205Lys Ser Lys Val Ser Val Thr Cys Leu
Val Lys Asp Phe Tyr Pro Pro 210 215
220Asp Ile Ser Val Glu Trp Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly225
230 235 240Lys Tyr Ser Thr
Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe 245
250 255Leu Tyr Ser Lys Leu Ser Leu Glu Thr Ser
Arg Trp Gln Gln Val Glu 260 265
270Ser Phe Thr Cys Ala Val Met His Glu Ala Leu His Asn His Tyr Thr
275 280 285Lys Thr Asp Ile Ser Glu Ser
Leu Gly Lys 290 29546337PRTArtificial
SequenceSynthetic ssGLP1-S8/GLP1-3G_III_ VAReqIgG2 46Met Ala Val Leu Gly
Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1 5
10 15Val Leu Ser His Ser Glu Gly Thr Phe Thr Ser
Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly
35 40 45Gly Ala Gly Gly Gly Gly Gly Ser
Gly Gly Gly Ser Gly Gly Gly Ser 50 55
60Gly Gly Gly Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu65
70 75 80Leu Leu Gly Gly Pro
Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp 85
90 95Thr Leu Met Ile Ser Arg Thr Pro Val Val Thr
Cys Val Val Val Asn 100 105
110Leu Ser Asp Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn
115 120 125Thr Glu Val His Ser Ala Ile
Thr Lys Gln Arg Glu Ala Gln Phe Asn 130 135
140Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp
Trp145 150 155 160Leu Ser
Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro
165 170 175Gln Pro Ile Ser Arg Ala Ile
Ser Arg Gly Lys Gly Pro Ser Arg Val 180 185
190Pro Gln Val Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala
Lys Ser 195 200 205Lys Val Ser Val
Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile 210
215 220Ser Val Glu Trp Gln Ser Asn Arg Trp Pro Glu Leu
Glu Gly Lys Tyr225 230 235
240Ser Thr Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr
245 250 255Ser Lys Leu Ser Leu
Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe 260
265 270Thr Cys Ala Val Met His Glu Ala Leu His Asn His
Tyr Thr Lys Thr 275 280 285Asp Ile
Ser Glu Ser Leu Gly Lys Gly Gly Gly Gly Ser Gly Gly Gly 290
295 300Gly His Ala Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu305 310 315
320Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly
325 330
335Gly47298PRTArtificial SequenceSynthetic ssGLP1-S8_I_VAReqIgG2 47Met
Ala Val Leu Gly Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1
5 10 15Val Leu Ser His Ser Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val
Lys Gly 35 40 45Gly Ala Gly Gly
Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser 50 55
60Gly Gly Gly Ser Ser Val Pro Lys Pro Gln Cys Pro Pro
Tyr Thr His65 70 75
80Ser Lys Phe Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro
85 90 95Lys Asp Thr Leu Met Ile
Ser Arg Thr Pro Val Val Thr Cys Val Val 100
105 110Val Asn Leu Ser Asp Gln Tyr Pro Asp Val Gln Phe
Ser Trp Tyr Val 115 120 125Asp Asn
Thr Glu Val His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln 130
135 140Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu
Pro Ile Gln His Gln145 150 155
160Asp Trp Leu Ser Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly
165 170 175Val Pro Gln Pro
Ile Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser 180
185 190Arg Val Pro Gln Val Tyr Val Leu Pro Pro His
Pro Asp Glu Leu Ala 195 200 205Lys
Ser Lys Val Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro 210
215 220Asp Ile Ser Val Glu Trp Gln Ser Asn Arg
Trp Pro Glu Leu Glu Gly225 230 235
240Lys Tyr Ser Thr Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr
Phe 245 250 255Leu Tyr Ser
Lys Leu Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu 260
265 270Ser Phe Thr Cys Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr 275 280
285Lys Thr Asp Ile Ser Glu Ser Leu Gly Lys 290
29548757PRTArtificial SequenceSynthetic Feline glucagon-like peptide 1
receptor (GLP1R) 48Met Gly Leu Val Ala Pro Val Val Leu Leu His Gln Asp
Asp Glu Glu1 5 10 15His
Gly Gln Asp Glu Gly Pro Glu Asp Gly Ser Gly Tyr Leu Leu Gly 20
25 30Thr Leu Thr Arg Phe Ser Ser Asp
Phe Asp Ser Ala Pro Glu Val Ile 35 40
45Leu Ala Pro Asp Asp Gln Leu Gln Leu Pro His Pro Ser Ser Arg Glu
50 55 60Asn Phe Trp Ala Arg Thr Gly Leu
Cys Ala Glu Ser Phe Leu Leu Arg65 70 75
80Pro Val Gly Pro Val Gly Pro Val Met Gly Trp Ser Glu
Gly Phe His 85 90 95Lys
Arg Asn Ser Arg Gln Glu Phe Leu Arg Arg Arg Leu Phe Ala Gly
100 105 110Gly Leu Cys Ala Ala Ser Thr
Gln Glu Ser Arg Asn Arg Cys Ser Ser 115 120
125Arg Gly Cys Lys Ser Ser Pro Ala Asp Cys Pro Glu Leu Asp Arg
Thr 130 135 140Gln His Leu Gly Asn Ser
Val Gly Pro Ile Gln Ala Ala His Gln Glu145 150
155 160Leu Ala Leu Gly Ala Gly Gly Pro Gly Asp Glu
Cys Gln Cys Cys Ser 165 170
175Val Ser Asn Ser Leu Phe Ile Pro Glu Pro Gln Ser Thr Cys Pro Tyr
180 185 190Asn Gly Tyr Thr Ser Trp
Pro Leu Glu Gly Asn Leu Arg Val Ala Cys 195 200
205Ala Pro Pro Pro Pro Pro Pro Ala Arg Thr Leu Phe Gly Gly
Ser Arg 210 215 220Arg Gly Ala Val Asp
Lys Lys Ala Gly Gly Gly Asn Arg Ser Pro Gly225 230
235 240Gly Gly Ala Gly Thr Gly Glu Phe Gly Ala
Pro Gly Ala Gly Gly Gly 245 250
255Leu Gly Arg Arg Pro Glu Val Gly Ala Trp Thr Ala Ala Glu Gly Thr
260 265 270Asn Pro Ala Asp Leu
Ala Ser Ser Pro Pro Pro Pro Ser Thr Arg Pro 275
280 285Pro Ala Ala Pro Arg Pro Pro Cys Ala Asp Phe Cys
Ala Ala Ser Pro 290 295 300Gln Thr Thr
Phe Pro Thr Pro Ser Pro Arg Arg Pro Leu Pro Ala Ser305
310 315 320Gly Gly Ala Thr Val Ser Leu
Ser Glu Thr Val Gln Lys Trp Arg Glu 325
330 335Tyr Arg His Gln Cys Gln Arg Phe Leu Thr Glu Ala
Pro Pro Pro Ala 340 345 350Thr
Gly Leu Phe Cys Asn Arg Thr Phe Asp Glu Tyr Ala Cys Trp Pro 355
360 365Asp Gly Leu Pro Gly Ser Phe Val Asn
Val Ser Cys Pro Trp Tyr Leu 370 375
380Pro Trp Ala Ser Ser Val Leu Gln Gly His Val Tyr Arg Phe Cys Thr385
390 395 400Ala Glu Gly Leu
Trp Leu Arg Gln Asp Asn Ser Ser Leu Pro Trp Arg 405
410 415Asn Leu Ser Glu Cys Glu Glu Ser Lys Arg
Gly Glu Arg Ser Ser Pro 420 425
430Glu Glu Gln Leu Leu Ser Phe Ser Ile Ile Tyr Thr Val Gly Tyr Thr
435 440 445Leu Ser Phe Ser Ala Leu Val
Ile Ala Ser Ala Ile Leu Leu Ser Phe 450 455
460Arg His Leu His Cys Thr Arg Asn Tyr Ile His Leu Asn Leu Phe
Ala465 470 475 480Ser Phe
Ile Leu Arg Ala Leu Ser Val Phe Ile Arg Asp Ala Val Leu
485 490 495Lys Trp Met Tyr Ser Thr Ala
Pro Gln Gln His Gln Trp Asp Gly Leu 500 505
510Leu Ser Tyr Gln Asp Ser Leu Gly Cys Arg Leu Val Phe Leu
Leu Met 515 520 525Gln Tyr Cys Val
Ala Ala Asn Tyr Tyr Trp Leu Leu Val Glu Gly Val 530
535 540Tyr Leu Tyr Thr Leu Leu Ala Phe Ser Val Phe Ser
Glu Gln Arg Ile545 550 555
560Phe Arg Leu Tyr Leu Ser Ile Gly Trp Gly Val Pro Leu Leu Phe Val
565 570 575Ile Trp Gly Ile Val
Lys Tyr Leu Tyr Glu Asp Glu Gly Cys Trp Thr 580
585 590Arg Asn Ser Asn Met Asn Tyr Trp Leu Ile Ile Arg
Leu Pro Ile Leu 595 600 605Phe Ala
Ile Gly Val Asn Phe Leu Ile Phe Val Arg Val Ile Cys Ile 610
615 620Val Val Ser Lys Leu Lys Ala Asn Leu Met Cys
Lys Thr Asp Ile Lys625 630 635
640Cys Arg Leu Ala Lys Ser Thr Leu Thr Leu Ile Pro Leu Leu Gly Thr
645 650 655His Glu Val Val
Phe Ala Phe Val Met Asp Glu His Ala Arg Gly Thr 660
665 670Leu Arg Phe Ile Lys Leu Phe Thr Glu Leu Ser
Phe Thr Ser Phe Gln 675 680 685Gly
Leu Met Val Ala Ile Leu Tyr Cys Phe Val Asn Asn Glu Val Gln 690
695 700Met Glu Phe Arg Arg Ser Trp Glu Arg Trp
Arg Leu Lys His Leu His705 710 715
720Ile Gln Arg Asp Ser Ser Met Lys Pro Leu Lys Cys Pro Thr Ser
Ser 725 730 735Leu Thr Ser
Gly Gly Thr Val Gly Ser Ser Val Tyr Ala Ala Ser Cys 740
745 750Gln Ala Ser Cys Ser
75549126PRTArtificial SequenceSynthetic Mature feline glucagon-like
peptide 1 receptor-N-terminal domain (GLP1R-N) 49Arg Pro Leu Pro Ala
Ser Gly Gly Ala Thr Val Ser Leu Ser Glu Thr1 5
10 15Val Gln Lys Trp Arg Glu Tyr Arg His Gln Cys
Gln Arg Phe Leu Thr 20 25
30Glu Ala Pro Pro Pro Ala Thr Gly Leu Phe Cys Asn Arg Thr Phe Asp
35 40 45Glu Tyr Ala Cys Trp Pro Asp Gly
Leu Pro Gly Ser Phe Val Asn Val 50 55
60Ser Cys Pro Trp Tyr Leu Pro Trp Ala Ser Ser Val Leu Gln Gly His65
70 75 80Val Tyr Arg Phe Cys
Thr Ala Glu Gly Leu Trp Leu Arg Gln Asp Asn 85
90 95Ser Ser Leu Pro Trp Arg Asn Leu Ser Glu Cys
Glu Glu Ser Lys Arg 100 105
110Gly Glu Arg Ser Ser Pro Glu Glu Gln Leu Leu Ser Phe Ser 115
120 12550392PRTArtificial SequenceSynthetic
ssFeGLP1R-N_huFc_ PolyHis 50Met Glu Thr Asp Thr Leu Leu Leu Trp Val Leu
Leu Leu Trp Val Pro1 5 10
15Gly Ser Thr Gly Arg Pro Leu Pro Ala Ser Gly Gly Ala Thr Val Ser
20 25 30Leu Ser Glu Thr Val Gln Lys
Trp Arg Glu Tyr Arg His Gln Cys Gln 35 40
45Arg Phe Leu Thr Glu Ala Pro Pro Pro Ala Thr Gly Leu Phe Cys
Asn 50 55 60Arg Thr Phe Asp Glu Tyr
Ala Cys Trp Pro Asp Gly Leu Pro Gly Ser65 70
75 80Phe Val Asn Val Ser Cys Pro Trp Tyr Leu Pro
Trp Ala Ser Ser Val 85 90
95Leu Gln Gly His Val Tyr Arg Phe Cys Thr Ala Glu Gly Leu Trp Leu
100 105 110Arg Gln Asp Asn Ser Ser
Leu Pro Trp Arg Asn Leu Ser Glu Cys Glu 115 120
125Glu Ser Lys Arg Gly Glu Arg Ser Ser Pro Glu Glu Gln Leu
Leu Ser 130 135 140Phe Ser Gly Ser Glu
Asn Leu Tyr Phe Gln Gly Pro Lys Ser Cys Asp145 150
155 160Lys Thr His Thr Cys Pro Pro Cys Pro Ala
Pro Glu Leu Leu Gly Gly 165 170
175Pro Ser Val Phe Leu Phe Pro Pro Lys Pro Lys Asp Thr Leu Met Ile
180 185 190Ser Arg Thr Pro Glu
Val Thr Cys Val Val Val Asp Val Ser His Glu 195
200 205Asp Pro Glu Val Lys Phe Asn Trp Tyr Val Asp Gly
Val Glu Val His 210 215 220Asn Ala Lys
Thr Lys Pro Arg Glu Glu Gln Tyr Asn Ser Thr Tyr Arg225
230 235 240Val Val Ser Val Leu Thr Val
Leu His Gln Asp Trp Leu Asn Gly Lys 245
250 255Glu Tyr Lys Cys Lys Val Ser Asn Lys Ala Leu Pro
Ala Pro Ile Glu 260 265 270Lys
Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro Gln Val Tyr 275
280 285Thr Leu Pro Pro Ser Arg Asp Glu Leu
Thr Lys Asn Gln Val Ser Leu 290 295
300Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp Ile Ala Val Glu Trp305
310 315 320Glu Ser Asn Gly
Gln Pro Glu Asn Asn Tyr Lys Thr Thr Pro Pro Val 325
330 335Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr
Ser Lys Leu Thr Val Asp 340 345
350Lys Ser Arg Trp Gln Gln Gly Asn Val Phe Ser Cys Ser Val Met His
355 360 365Glu Ala Leu His Asn His Tyr
Thr Gln Lys Ser Leu Ser Leu Ser Pro 370 375
380Gly Lys His His His His His His385
39051156PRTArtificial SequenceSynthetic ssFeGLP1R-N_PolyHis 51Met Glu Thr
Asp Thr Leu Leu Leu Trp Val Leu Leu Leu Trp Val Pro1 5
10 15Gly Ser Thr Gly Arg Pro Leu Pro Ala
Ser Gly Gly Ala Thr Val Ser 20 25
30Leu Ser Glu Thr Val Gln Lys Trp Arg Glu Tyr Arg His Gln Cys Gln
35 40 45Arg Phe Leu Thr Glu Ala Pro
Pro Pro Ala Thr Gly Leu Phe Cys Asn 50 55
60Arg Thr Phe Asp Glu Tyr Ala Cys Trp Pro Asp Gly Leu Pro Gly Ser65
70 75 80Phe Val Asn Val
Ser Cys Pro Trp Tyr Leu Pro Trp Ala Ser Ser Val 85
90 95Leu Gln Gly His Val Tyr Arg Phe Cys Thr
Ala Glu Gly Leu Trp Leu 100 105
110Arg Gln Asp Asn Ser Ser Leu Pro Trp Arg Asn Leu Ser Glu Cys Glu
115 120 125Glu Ser Lys Arg Gly Glu Arg
Ser Ser Pro Glu Glu Gln Leu Leu Ser 130 135
140Phe Ser Gly Gly Gly Ser His His His His His His145
150 15552326PRTArtificial SequenceSynthetic
GLP1-G8/Gluc-3G_IV_ WTfeIgG2 52His Gly Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly Gly
20 25 30Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser 35 40
45Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly
Pro Lys 50 55 60Cys Pro Val Pro Glu
Ile Pro Gly Ala Pro Ser Val Phe Ile Phe Pro65 70
75 80Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser
Arg Thr Pro Glu Val Thr 85 90
95Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr
100 105 110Trp Phe Val Asp Asn
Thr Glu Met His Thr Ala Lys Thr Arg Pro Arg 115
120 125Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser
Val Leu Pro Ile 130 135 140Leu His Gln
Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn145
150 155 160Ser Lys Ser Leu Pro Ser Ala
Met Glu Arg Thr Ile Ser Lys Ala Lys 165
170 175Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro
Pro Thr Gln Glu 180 185 190Glu
Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Gly Phe 195
200 205His Pro Pro Asp Ile Ala Val Glu Trp
Glu Ile Thr Gly Gln Pro Glu 210 215
220Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly225
230 235 240Thr Tyr Phe Leu
Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp Gln 245
250 255Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser
His Glu Ala Leu His Ser 260 265
270His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys Gly Gly Gly
275 280 285Gly Ser Gly Gly Gly Gly His
Ser Gln Gly Thr Phe Thr Ser Asp Tyr 290 295
300Ser Lys Tyr Leu Asp Ser Arg Arg Ala Gln Asp Phe Val Gln Trp
Leu305 310 315 320Met Asn
Thr Gly Gly Gly 32553326PRTArtificial SequenceSynthetic
Gluc/GLP1-2G_V_ WTfeIgG2 53His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser
Lys Tyr Leu Asp Ser1 5 10
15Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Gly Gly Gly
20 25 30Gly Ser Gly Gly Gly Gly Ser
Gly Gly Gly Gly Ser Gly Gly Gly Gly 35 40
45Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly
Pro 50 55 60Lys Cys Pro Val Pro Glu
Ile Pro Gly Ala Pro Ser Val Phe Ile Phe65 70
75 80Pro Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser
Arg Thr Pro Glu Val 85 90
95Thr Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile
100 105 110Thr Trp Phe Val Asp Asn
Thr Glu Met His Thr Ala Lys Thr Arg Pro 115 120
125Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val
Leu Pro 130 135 140Ile Leu His Gln Asp
Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val145 150
155 160Asn Ser Lys Ser Leu Pro Ser Ala Met Glu
Arg Thr Ile Ser Lys Ala 165 170
175Lys Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln
180 185 190Glu Glu Leu Ser Glu
Asn Lys Val Ser Val Thr Cys Leu Ile Lys Gly 195
200 205Phe His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile
Thr Gly Gln Pro 210 215 220Glu Pro Glu
Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp225
230 235 240Gly Thr Tyr Phe Leu Tyr Ser
Arg Leu Ser Val Asp Arg Ser His Trp 245
250 255Gln Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His
Glu Ala Leu His 260 265 270Ser
His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys Gly Gly 275
280 285Gly Gly Ser Gly Gly Gly Gly His Ala
Glu Gly Thr Phe Thr Ser Asp 290 295
300Val Ser Ser Tyr Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp305
310 315 320Leu Val Lys Gly
Gly Gly 32554324PRTArtificial SequenceSynthetic
GLP1-G8/Glu-4G_IV_ VARcaIgGD 54His Gly Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Pro Lys Glu Ser Thr Ser Lys Cys Ile Ser Pro Cys Pro
Val Pro 50 55 60Glu Ser Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys65 70
75 80Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu
Ile Thr Cys Val Val Leu 85 90
95Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp Phe Val Asp
100 105 110Gly Lys Glu Val His
Thr Ala Lys Thr Gln Pro Arg Glu Gln Gln Phe 115
120 125Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
Gly His Gln Asp 130 135 140Trp Leu Thr
Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile Gly Leu145
150 155 160Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly Gln Ala His 165
170 175Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys
Glu Leu Ser Ser 180 185 190Ser
Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe Pro Pro 195
200 205Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Pro Glu Pro Glu Ser 210 215
220Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly Ser Tyr Phe225
230 235 240Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly Asp 245
250 255Thr Phe Thr Cys Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr 260 265
270Asp Leu Ser Leu Ser His Ser Pro Gly Lys Gly Gly Gly Gly Ser Gly
275 280 285Gly Gly Gly His Ser Gln Gly
Thr Phe Thr Ser Asp Tyr Ser Lys Tyr 290 295
300Leu Asp Ser Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn
Thr305 310 315 320Gly Gly
Gly Gly55323PRTArtificial SequenceSynthetic Gluc/GLP1-3G_V_ VARcaIgGD
55His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser1
5 10 15Arg Arg Ala Gln Asp Phe
Val Gln Trp Leu Met Asn Thr Gly Ala Gly 20 25
30Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser
Gly Gly Gly 35 40 45Ser Pro Lys
Glu Ser Thr Ser Lys Cys Ile Ser Pro Cys Pro Val Pro 50
55 60Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro Lys Pro Lys65 70 75
80Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Ile Thr Cys Val Val Leu
85 90 95Asp Leu Gly Arg Glu Asp
Pro Glu Val Gln Ile Ser Trp Phe Val Asp 100
105 110Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg
Glu Gln Gln Phe 115 120 125Asn Ser
Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly His Gln Asp 130
135 140Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val
Asn His Ile Gly Leu145 150 155
160Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly Gln Ala His
165 170 175Gln Pro Ser Val
Tyr Val Leu Pro Pro Ser Pro Lys Glu Leu Ser Ser 180
185 190Ser Asp Thr Val Thr Leu Thr Cys Leu Ile Lys
Asp Phe Phe Pro Pro 195 200 205Glu
Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro Glu Pro Glu Ser 210
215 220Lys Tyr His Thr Thr Ala Pro Gln Leu Asp
Glu Asp Gly Ser Tyr Phe225 230 235
240Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly
Asp 245 250 255Thr Phe Thr
Cys Ala Val Met His Glu Ala Leu His Asn His Tyr Thr 260
265 270Asp Leu Ser Leu Ser His Ser Pro Gly Lys
Gly Gly Gly Gly Ser Gly 275 280
285Gly Gly Gly His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr 290
295 300Leu Glu Gly Gln Ala Ala Lys Glu
Phe Ile Ala Trp Leu Val Lys Gly305 310
315 320Gly Gly Gly56319PRTArtificial SequenceSynthetic
GLP1-G8/Gluc-4G_IV_ VAReqIgGD 56His Gly Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly
Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu
Leu Gly 50 55 60Gly Pro Ser Val Phe
Ile Phe Pro Pro Asn Pro Lys Asp Thr Leu Met65 70
75 80Ile Ser Arg Thr Pro Val Val Thr Cys Val
Val Val Asn Leu Ser Asp 85 90
95Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val
100 105 110His Ser Ala Ile Thr
Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr 115
120 125Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp
Trp Leu Ser Gly 130 135 140Lys Glu Phe
Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile145
150 155 160Ser Arg Ala Ile Ser Arg Gly
Lys Gly Pro Ser Arg Val Pro Gln Val 165
170 175Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys
Ser Lys Val Ser 180 185 190Val
Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu 195
200 205Trp Gln Ser Asn Arg Trp Pro Glu Leu
Glu Gly Lys Tyr Ser Thr Thr 210 215
220Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu225
230 235 240Ser Leu Glu Thr
Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala 245
250 255Val Met His Glu Ala Leu His Asn His Tyr
Thr Lys Thr Asp Ile Ser 260 265
270Glu Ser Leu Gly Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly His Ser
275 280 285Gln Gly Thr Phe Thr Ser Asp
Tyr Ser Lys Tyr Leu Asp Ser Arg Arg 290 295
300Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Gly Gly Gly Gly305
310 31557318PRTArtificial SequenceSynthetic
Gluc/GLP1-3G_V_ VAReqIgG2 57His Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser
Lys Tyr Leu Asp Ser1 5 10
15Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly Gly
Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu Leu
Gly 50 55 60Gly Pro Ser Val Phe Ile
Phe Pro Pro Asn Pro Lys Asp Thr Leu Met65 70
75 80Ile Ser Arg Thr Pro Val Val Thr Cys Val Val
Val Asn Leu Ser Asp 85 90
95Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val
100 105 110His Ser Ala Ile Thr Lys
Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr 115 120
125Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu
Ser Gly 130 135 140Lys Glu Phe Lys Cys
Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile145 150
155 160Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro
Ser Arg Val Pro Gln Val 165 170
175Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
180 185 190Val Thr Cys Leu Val
Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu 195
200 205Trp Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys
Tyr Ser Thr Thr 210 215 220Pro Ala Gln
Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys Leu225
230 235 240Ser Leu Glu Thr Ser Arg Trp
Gln Gln Val Glu Ser Phe Thr Cys Ala 245
250 255Val Met His Glu Ala Leu His Asn His Tyr Thr Lys
Thr Asp Ile Ser 260 265 270Glu
Ser Leu Gly Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly His Ala 275
280 285Glu Gly Thr Phe Thr Ser Asp Val Ser
Ser Tyr Leu Glu Gly Gln Ala 290 295
300Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly Gly305
310 31558321PRTArtificial SequenceSynthetic
GLP1-G8/Glu-4G_IV_ huIgG4 58His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser
Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Ala Gly
20 25 30Gly Gly Gly Gly Ser Gly Gly
Gly Ser Gly Gly Gly Ser Gly Gly Gly 35 40
45Ser Ser Glu Ser Lys Tyr Gly Pro Pro Cys Pro Pro Cys Pro Ala
Pro 50 55 60Glu Phe Leu Gly Gly Pro
Ser Val Phe Leu Phe Pro Pro Lys Pro Lys65 70
75 80Asp Thr Leu Met Ile Ser Arg Thr Pro Glu Val
Thr Cys Val Val Val 85 90
95Asp Val Ser Gln Glu Asp Pro Glu Val Gln Phe Asn Trp Tyr Val Asp
100 105 110Gly Val Glu Val His Asn
Ala Lys Thr Lys Pro Arg Glu Glu Gln Phe 115 120
125Asn Ser Thr Tyr Arg Val Val Ser Val Leu Thr Val Leu His
Gln Asp 130 135 140Trp Leu Asn Gly Lys
Glu Tyr Lys Cys Lys Val Ser Asn Lys Gly Leu145 150
155 160Pro Ser Ser Ile Glu Lys Thr Ile Ser Lys
Ala Lys Gly Gln Pro Arg 165 170
175Glu Pro Gln Val Tyr Thr Leu Pro Pro Ser Gln Glu Glu Met Thr Lys
180 185 190Asn Gln Val Ser Leu
Thr Cys Leu Val Lys Gly Phe Tyr Pro Ser Asp 195
200 205Ile Ala Val Glu Trp Glu Ser Asn Gly Gln Pro Glu
Asn Asn Tyr Lys 210 215 220Thr Thr Pro
Pro Val Leu Asp Ser Asp Gly Ser Phe Phe Leu Tyr Ser225
230 235 240Arg Leu Thr Val Asp Lys Ser
Arg Trp Gln Glu Gly Asn Val Phe Ser 245
250 255Cys Ser Val Met His Glu Ala Leu His Asn His Tyr
Thr Gln Lys Ser 260 265 270Leu
Ser Leu Ser Leu Gly Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly 275
280 285His Ser Gln Gly Thr Phe Thr Ser Asp
Tyr Ser Lys Tyr Leu Asp Ser 290 295
300Arg Arg Ala Gln Asp Phe Val Gln Trp Leu Met Asn Thr Gly Gly Gly305
310 315
320Gly59321PRTArtificial SequenceSynthetic Gluc/GLP1-3G_V_huIgG4 59His
Ser Gln Gly Thr Phe Thr Ser Asp Tyr Ser Lys Tyr Leu Asp Ser1
5 10 15Arg Arg Ala Gln Asp Phe Val
Gln Trp Leu Met Asn Thr Gly Gly Gly 20 25
30Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
Ser Ser 35 40 45Glu Ser Lys Tyr
Gly Pro Pro Cys Pro Pro Cys Pro Ala Pro Glu Phe 50 55
60Leu Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Lys Pro
Lys Asp Thr65 70 75
80Leu Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val
85 90 95Ser Gln Glu Asp Pro Glu
Val Gln Phe Asn Trp Tyr Val Asp Gly Val 100
105 110Glu Val His Asn Ala Lys Thr Lys Pro Arg Glu Glu
Gln Phe Asn Ser 115 120 125Thr Tyr
Arg Val Val Ser Val Leu Thr Val Leu His Gln Asp Trp Leu 130
135 140Asn Gly Lys Glu Tyr Lys Cys Lys Val Ser Asn
Lys Gly Leu Pro Ser145 150 155
160Ser Ile Glu Lys Thr Ile Ser Lys Ala Lys Gly Gln Pro Arg Glu Pro
165 170 175Gln Val Tyr Thr
Leu Pro Pro Ser Gln Glu Glu Met Thr Lys Asn Gln 180
185 190Val Ser Leu Thr Cys Leu Val Lys Gly Phe Tyr
Pro Ser Asp Ile Ala 195 200 205Val
Glu Trp Glu Ser Asn Gly Gln Pro Glu Asn Asn Tyr Lys Thr Thr 210
215 220Pro Pro Val Leu Asp Ser Asp Gly Ser Phe
Phe Leu Tyr Ser Arg Leu225 230 235
240Thr Val Asp Lys Ser Arg Trp Gln Glu Gly Asn Val Phe Ser Cys
Ser 245 250 255Val Met His
Glu Ala Leu His Asn His Tyr Thr Gln Lys Ser Leu Ser 260
265 270Leu Ser Leu Gly Lys Gly Gly Gly Gly Ser
Gly Gly Gly Gly His Ala 275 280
285Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly Gln Ala 290
295 300Ala Lys Glu Phe Ile Ala Trp Leu
Val Lys Gly Gly Ala Gly Gly Gly305 310
315 320Gly60221PRTArtificial SequenceSynthetic Variant
canine IgG-A Fc , C1q -, Protein A +, I(21)T, Q(207)H 60Pro Val Pro
Glu Pro Leu Gly Gly Pro Ser Val Leu Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Thr Leu Arg Ile Thr
Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Glu Val
His Thr Ala Lys Thr Gln Ser Arg Glu 50 55
60Gln Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65
70 75 80His Gln Asp Trp
Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His 85
90 95Ile Asp Leu Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly 100 105
110Arg Ala His Lys Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu
115 120 125Leu Ser Ser Ser Asp Thr Val
Ser Ile Thr Cys Leu Ile Lys Asp Phe 130 135
140Tyr Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln
Glu145 150 155 160Pro Glu
Arg Lys His Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln 180 185
190Gln Gly Asp Pro Phe Thr Cys Ala Val Met His Glu Thr Leu
His Asn 195 200 205His Tyr Thr Asp
Leu Ser Leu Ser His Ser Pro Gly Lys 210 215
22061220PRTArtificial SequenceSynthetic Variant canine IgG-C Fc ,
C1q +, Protein A +, I(21)T 61Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser
Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp
Pro Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro
Arg Glu 50 55 60Glu Gln Ser Asn Gly
Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe
Lys Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro
Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val
Lys Asp Phe Phe 130 135 140Pro Pro Glu
Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Met Thr
Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser
Pro Gly Lys 210 215
22062221PRTArtificial SequenceSynthetic Variant canine IgG-D Fc , C1q -,
Protein A +, I(21)T, Q(207)H 62Pro Val Pro Glu Ser Leu Gly Gly Pro
Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Arg Ile Thr Arg Thr Pro Glu Ile Thr
Cys 20 25 30Val Val Leu Asp
Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr
Gln Pro Arg Glu 50 55 60Gln Gln Phe
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65 70
75 80His Gln Asp Trp Leu Thr Gly Lys
Glu Phe Lys Cys Arg Val Asn His 85 90
95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala
Arg Gly 100 105 110Gln Ala His
Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu 115
120 125Leu Ser Ser Ser Asp Thr Val Thr Leu Thr Cys
Leu Ile Lys Asp Phe 130 135 140Phe Pro
Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro Glu145
150 155 160Pro Glu Ser Lys Tyr His Thr
Thr Ala Pro Gln Leu Asp Glu Asp Gly 165
170 175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys
Ser Arg Trp Gln 180 185 190Gln
Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu His Asn 195
200 205His Tyr Thr Asp Leu Ser Leu Ser His
Ser Pro Gly Lys 210 215
22063214PRTUnknownWild-type equine IgG1 Fc , Protein A +, C1q + 63Gly Gly
Pro Ser Val Phe Leu Phe Pro Pro Asn Pro Lys Asp Thr Leu1 5
10 15Met Ile Thr Arg Thr Pro Glu Val
Thr Cys Val Val Val Asp Val Ser 20 25
30Gln Glu Asn Pro Asp Val Lys Phe Asn Trp Tyr Met Asp Gly Val
Glu 35 40 45Val Arg Thr Ala Thr
Thr Arg Pro Lys Glu Glu Gln Phe Asn Ser Thr 50 55
60Tyr Arg Val Val Ser Val Leu Arg Ile Gln His Gln Asp Trp
Leu Ser65 70 75 80Gly
Lys Glu Phe Lys Cys Lys Val Asn Asn Gln Ala Leu Pro Gln Pro
85 90 95Ile Glu Arg Thr Ile Thr Lys
Thr Lys Gly Arg Ser Gln Glu Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Pro Asp Glu Ser Lys Lys Ser
Lys Val 115 120 125Ser Val Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Glu Ile Asn Ile 130
135 140Glu Trp Gln Ser Asn Gly Gln Pro Glu Leu Glu Thr
Lys Tyr Ser Thr145 150 155
160Thr Gln Ala Gln Gln Asp Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Ser Val Asp Arg
Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys 180
185 190Gly Val Met His Glu Ala Leu His Asn His Tyr Thr
Gln Lys Asn Val 195 200 205Ser Lys
Asn Pro Gly Lys 21064214PRTUnknownWild-type equine IgG2 Fc, Protein A
-, C1q - 64Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala
Leu1 5 10 15Met Ile Ser
Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser 20
25 30Asp Gln Tyr Pro Asp Val Gln Phe Ser Trp
Tyr Val Asp Asn Thr Glu 35 40
45Val His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro
Gln Pro 85 90 95Ile Ser
Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln 100
105 110Val Tyr Val Leu Pro Pro His Pro Asp
Glu Leu Ala Lys Ser Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val
130 135 140Glu Trp Gln Ser Asn Arg Trp
Pro Glu Leu Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys
180 185 190Ala Val Met His Glu Ala
Leu His Asn His Phe Thr Lys Thr Asp Ile 195 200
205Ser Glu Ser Leu Gly Lys 21065214PRTUnknownWild-type
equine IgG3 Fc, Protein A +, C1q+ 65Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro Lys Pro Lys Asp Val Leu1 5 10
15Met Ile Thr Arg Met Pro Glu Val Thr Cys Leu Val Val Asp Val
Ser 20 25 30His Asp Ser Ser
Asp Val Leu Phe Thr Trp Tyr Val Asp Gly Thr Glu 35
40 45Val Lys Thr Ala Lys Thr Met Pro Asn Glu Glu Gln
Asn Asn Ser Thr 50 55 60Tyr Arg Val
Val Ser Val Leu Arg Ile Gln His Gln Asp Trp Leu Asn65 70
75 80Gly Lys Lys Phe Lys Cys Lys Val
Asn Asn Gln Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Ser Lys Ala Thr Gly Gln Thr Arg Val
Pro Gln 100 105 110Val Tyr Val
Leu Ala Pro His Pro Asp Glu Leu Ser Lys Asn Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro
Pro Asp Ile Thr Val 130 135 140Glu Trp
Gln Ser Asn Glu His Pro Glu Pro Glu Gly Lys Tyr Arg Thr145
150 155 160Thr Glu Ala Gln Lys Asp Ser
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Lys Asp Arg Trp Gln Gln Gly Thr
Thr Phe Thr Cys 180 185 190Val
Val Met His Glu Ala Leu His Asn His Val Met Gln Lys Asn Ile 195
200 205Ser Lys Asn Pro Gly Lys
21066214PRTUnknownWild-type equine IgG4 Fc, Protein A +, C1q + 66Val Gly
Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1 5
10 15Met Ile Ser Arg Thr Pro Thr Val
Thr Cys Val Val Val Asp Val Gly 20 25
30His Asp Phe Pro Asp Val Gln Phe Asn Trp Tyr Val Asp Gly Val
Glu 35 40 45Thr His Thr Ala Thr
Thr Glu Pro Lys Gln Glu Gln Phe Asn Ser Thr 50 55
60Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Lys Asp Trp
Leu Ser65 70 75 80Gly
Lys Glu Phe Lys Cys Lys Val Asn Asn Lys Ala Leu Pro Ala Pro
85 90 95Val Glu Arg Thr Ile Ser Ala
Pro Thr Gly Gln Pro Arg Glu Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Arg Asp Glu Leu Ser Lys Asn
Lys Val 115 120 125Ser Val Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Asp Ile 130
135 140Glu Trp Lys Ser Asn Gly Gln Pro Glu Pro Glu Thr
Lys Tyr Ser Thr145 150 155
160Thr Pro Ala Gln Leu Asp Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Thr Val Glu Thr
Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys 180
185 190Ala Val Met His Glu Ala Leu His Asn His Tyr Thr
Glu Lys Ser Val 195 200 205Ser Lys
Ser Pro Gly Lys 21067214PRTUnknownWild-type equine IgG5 Fc, Protein A
-, C1q - 67Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val
Leu1 5 10 15Met Ile Ser
Arg Lys Pro Glu Val Thr Cys Val Val Val Asp Leu Gly 20
25 30His Asp Asp Pro Asp Val Gln Phe Thr Trp
Phe Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Glu Glu Gln Phe Asn Ser Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Thr Ser Lys Ala Leu Pro
Ala Pro 85 90 95Val Glu
Arg Thr Ile Ser Lys Ala Lys Gly Gln Leu Arg Val Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Pro Asp
Glu Leu Ala Lys Asn Thr Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Glu Ile Asp Val
130 135 140Glu Trp Gln Ser Asn Glu His
Pro Glu Pro Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asn Ser Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Ser Val Glu Thr Ser Arg Trp Lys Gln Gly Glu Ser Phe Thr Cys
180 185 190Gly Val Met His Glu Ala
Val Glu Asn His Tyr Thr Gln Lys Asn Val 195 200
205Ser His Ser Pro Gly Lys 21068214PRTUnknownWild-type
equine IgG6 Fc, Protein A -, C1q - 68Gly Arg Pro Ser Val Phe Ile Phe Pro
Pro Asn Pro Lys Asp Thr Leu1 5 10
15Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val
Ser 20 25 30Gln Glu Asn Pro
Asp Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 35
40 45Ala His Thr Ala Thr Thr Lys Ala Lys Glu Lys Gln
Asp Asn Ser Thr 50 55 60Tyr Arg Val
Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Arg Arg65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val
Asn Asn Arg Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Thr Lys Ala Lys Gly Glu Leu Gln Asp
Pro Gln 100 105 110Val Tyr Ile
Leu Ala Pro His Pro Asp Glu Val Thr Lys Asn Thr Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro
Pro Asp Ile Asn Val 130 135 140Glu Trp
Gln Ser Asn Glu Glu Pro Glu Pro Glu Val Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asp Gly
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Thr Asp Arg Trp Glu Gln Gly Glu
Ser Phe Thr Cys 180 185 190Val
Val Met His Glu Ala Ile Arg His Thr Tyr Arg Gln Lys Ser Ile 195
200 205Thr Asn Phe Pro Gly Lys
21069214PRTUnknownWild-type equine IgG7 Fc, Protein A +, C1q + 69Val Gly
Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1 5
10 15Met Ile Ser Arg Thr Pro Thr Val
Thr Cys Val Val Val Asp Val Gly 20 25
30His Asp Phe Pro Asp Val Gln Phe Asn Trp Tyr Val Asp Gly Val
Glu 35 40 45Thr His Thr Ala Thr
Thr Glu Pro Lys Gln Glu Gln Asn Asn Ser Thr 50 55
60Tyr Arg Val Val Ser Ile Leu Ala Ile Gln His Lys Asp Trp
Leu Ser65 70 75 80Gly
Lys Glu Phe Lys Cys Lys Val Asn Asn Gln Ala Leu Pro Ala Pro
85 90 95Val Gln Lys Thr Ile Ser Lys
Pro Thr Gly Gln Pro Arg Glu Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Pro Asp Glu Leu Ser Lys Asn
Lys Val 115 120 125Ser Val Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Asp Ile 130
135 140Glu Trp Lys Ser Asn Gly Gln Pro Glu Pro Glu Thr
Lys Tyr Ser Thr145 150 155
160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Thr Val Glu Thr
Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys 180
185 190Ala Val Met His Glu Ala Leu His Asn His Tyr Thr
Glu Lys Ser Val 195 200 205Ser Lys
Ser Pro Gly Lys 21070214PRTArtificial SequenceSynthetic Variant equine
IgG1 Fc , Protein A +, C1q - , K(87)S 70Gly Gly Pro Ser Val Phe Leu
Phe Pro Pro Asn Pro Lys Asp Thr Leu1 5 10
15Met Ile Thr Arg Thr Pro Glu Val Thr Cys Val Val Val
Asp Val Ser 20 25 30Gln Glu
Asn Pro Asp Val Lys Phe Asn Trp Tyr Met Asp Gly Val Glu 35
40 45Val Arg Thr Ala Thr Thr Arg Pro Lys Glu
Glu Gln Phe Asn Ser Thr 50 55 60Tyr
Arg Val Val Ser Val Leu Arg Ile Gln His Gln Asp Trp Leu Ser65
70 75 80Gly Lys Glu Phe Lys Cys
Ser Val Asn Asn Gln Ala Leu Pro Gln Pro 85
90 95Ile Glu Arg Thr Ile Thr Lys Thr Lys Gly Arg Ser
Gln Glu Pro Gln 100 105 110Val
Tyr Val Leu Ala Pro His Pro Asp Glu Ser Lys Lys Ser Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe
Tyr Pro Pro Glu Ile Asn Ile 130 135
140Glu Trp Gln Ser Asn Gly Gln Pro Glu Leu Glu Thr Lys Tyr Ser Thr145
150 155 160Thr Gln Ala Gln
Gln Asp Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Ser Val Asp Arg Asn Arg Trp Gln Gln
Gly Thr Thr Phe Thr Cys 180 185
190Gly Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Asn Val
195 200 205Ser Lys Asn Pro Gly Lys
21071214PRTArtificial SequenceSynthetic Variant equine IgG2 Fc, C1q -,
Protein A +, F(203)Y 71Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro
Lys Asp Ala Leu1 5 10
15Met Ile Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser
20 25 30Asp Gln Tyr Pro Asp Val Gln
Phe Ser Trp Tyr Val Asp Asn Thr Glu 35 40
45Val His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser
Thr 50 55 60Tyr Arg Val Val Ser Val
Leu Pro Ile Gln His Gln Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val
Gly Val Pro Gln Pro 85 90
95Ile Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln
100 105 110Val Tyr Val Leu Pro Pro
His Pro Asp Glu Leu Ala Lys Ser Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile
Ser Val 130 135 140Glu Trp Gln Ser Asn
Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser
Tyr Phe Leu Tyr Ser Lys 165 170
175Leu Ser Leu Glu Thr Ser Arg Trp Gln Gln Gly Glu Ser Phe Thr Cys
180 185 190Ala Val Met His Glu
Ala Leu His Asn His Tyr Thr Lys Thr Asp Ile 195
200 205Ser Glu Ser Leu Gly Lys 21072214PRTArtificial
SequenceSynthetic Variant equine IgG2 Fc, C1q -, Protein A +,
A(15)T, F(203)Y 72Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp
Thr Leu1 5 10 15Met Ile
Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser 20
25 30Asp Gln Tyr Pro Asp Val Gln Phe Ser
Trp Tyr Val Asp Asn Thr Glu 35 40
45Val His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro
Gln Pro 85 90 95Ile Ser
Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln 100
105 110Val Tyr Val Leu Pro Pro His Pro Asp
Glu Leu Ala Lys Ser Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val
130 135 140Glu Trp Gln Ser Asn Arg Trp
Pro Glu Leu Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys
180 185 190Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr Lys Thr Asp Ile 195 200
205Ser Glu Ser Leu Gly Lys 21073214PRTArtificial
SequenceSynthetic Variant equine IgG3 Fc, Protein A +, C1q - ,
K(87)S 73Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1
5 10 15Met Ile Thr Arg
Met Pro Glu Val Thr Cys Leu Val Val Asp Val Ser 20
25 30His Asp Ser Ser Asp Val Leu Phe Thr Trp Tyr
Val Asp Gly Thr Glu 35 40 45Val
Lys Thr Ala Lys Thr Met Pro Asn Glu Glu Gln Asn Asn Ser Thr 50
55 60Tyr Arg Val Val Ser Val Leu Arg Ile Gln
His Gln Asp Trp Leu Asn65 70 75
80Gly Lys Lys Phe Lys Cys Ser Val Asn Asn Gln Ala Leu Pro Ala
Pro 85 90 95Val Glu Arg
Thr Ile Ser Lys Ala Thr Gly Gln Thr Arg Val Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Pro Asp Glu
Leu Ser Lys Asn Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Thr Val 130
135 140Glu Trp Gln Ser Asn Glu His Pro
Glu Pro Glu Gly Lys Tyr Arg Thr145 150
155 160Thr Glu Ala Gln Lys Asp Ser Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Lys Asp Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Val Val Met His Glu Ala
Leu His Asn His Val Met Gln Lys Asn Ile 195 200
205Ser Lys Asn Pro Gly Lys 21074214PRTArtificial
SequenceSynthetic Variant equine IgG4 Fc, Protein A +, C1q - ,
K(87)S 74Val Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1
5 10 15Met Ile Ser Arg
Thr Pro Thr Val Thr Cys Val Val Val Asp Val Gly 20
25 30His Asp Phe Pro Asp Val Gln Phe Asn Trp Tyr
Val Asp Gly Val Glu 35 40 45Thr
His Thr Ala Thr Thr Glu Pro Lys Gln Glu Gln Phe Asn Ser Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile Gln
His Lys Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Asn Asn Lys Ala Leu Pro Ala
Pro 85 90 95Val Glu Arg
Thr Ile Ser Ala Pro Thr Gly Gln Pro Arg Glu Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Arg Asp Glu
Leu Ser Lys Asn Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Asp Ile 130
135 140Glu Trp Lys Ser Asn Gly Gln Pro
Glu Pro Glu Thr Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Ser Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Thr Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr Glu Lys Ser Val 195 200
205Ser Lys Ser Pro Gly Lys 21075214PRTArtificial
SequenceSynthetic Variant equine IgG5 Fc, C1q -, Protein A +,
V(199)L, E(200)H 75Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys
Asp Val Leu1 5 10 15Met
Ile Ser Arg Lys Pro Glu Val Thr Cys Val Val Val Asp Leu Gly 20
25 30His Asp Asp Pro Asp Val Gln Phe
Thr Trp Phe Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Glu Glu Gln Phe Asn Ser Thr
50 55 60Tyr Arg Val Val Ser Val Leu Pro
Ile Gln His Gln Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Thr Ser Lys Ala Leu
Pro Ala Pro 85 90 95Val
Glu Arg Thr Ile Ser Lys Ala Lys Gly Gln Leu Arg Val Pro Gln
100 105 110Val Tyr Val Leu Ala Pro His
Pro Asp Glu Leu Ala Lys Asn Thr Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Glu Ile Asp
Val 130 135 140Glu Trp Gln Ser Asn Glu
His Pro Glu Pro Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asn Ser Asp Gly Ser Tyr
Phe Leu Tyr Ser Lys 165 170
175Leu Ser Val Glu Thr Ser Arg Trp Lys Gln Gly Glu Ser Phe Thr Cys
180 185 190Gly Val Met His Glu Ala
Leu His Asn His Tyr Thr Gln Lys Asn Val 195 200
205Ser His Ser Pro Gly Lys 21076214PRTArtificial
SequenceSynthetic Variant equine IgG6 Fc, C1q -, Protein A +,
I(199)L , R(200)H, H(201)N, T(202)H 76Gly Arg Pro Ser Val Phe Ile Phe Pro
Pro Asn Pro Lys Asp Thr Leu1 5 10
15Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val
Ser 20 25 30Gln Glu Asn Pro
Asp Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 35
40 45Ala His Thr Ala Thr Thr Lys Ala Lys Glu Lys Gln
Asp Asn Ser Thr 50 55 60Tyr Arg Val
Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Arg Arg65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val
Asn Asn Arg Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Thr Lys Ala Lys Gly Glu Leu Gln Asp
Pro Gln 100 105 110Val Tyr Ile
Leu Ala Pro His Pro Asp Glu Val Thr Lys Asn Thr Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro
Pro Asp Ile Asn Val 130 135 140Glu Trp
Gln Ser Asn Glu Glu Pro Glu Pro Glu Val Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asp Gly
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Thr Asp Arg Trp Glu Gln Gly Glu
Ser Phe Thr Cys 180 185 190Val
Val Met His Glu Ala Leu His Asn His Tyr Arg Gln Lys Ser Ile 195
200 205Thr Asn Phe Pro Gly Lys
21077214PRTArtificial SequenceSynthetic Variant equine IgG7 Fc, Protein A
+, C1q - , K(87)S 77Val Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro
Lys Asp Val Leu1 5 10
15Met Ile Ser Arg Thr Pro Thr Val Thr Cys Val Val Val Asp Val Gly
20 25 30His Asp Phe Pro Asp Val Gln
Phe Asn Trp Tyr Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Gln Glu Gln Asn Asn Ser
Thr 50 55 60Tyr Arg Val Val Ser Ile
Leu Ala Ile Gln His Lys Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Ser Val Asn Asn Gln
Ala Leu Pro Ala Pro 85 90
95Val Gln Lys Thr Ile Ser Lys Pro Thr Gly Gln Pro Arg Glu Pro Gln
100 105 110Val Tyr Val Leu Ala Pro
His Pro Asp Glu Leu Ser Lys Asn Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile
Asp Ile 130 135 140Glu Trp Lys Ser Asn
Gly Gln Pro Glu Pro Glu Thr Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser
Tyr Phe Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Thr Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Ala Val Met His Glu
Ala Leu His Asn His Tyr Thr Glu Lys Ser Val 195
200 205Ser Lys Ser Pro Gly Lys 21078220PRTArtificial
SequenceSynthetic Variant canine IgG-B Fc, Protein A +, C1q - ,
K(93)R 78Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp
Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu
Val Gln Ile Ser Trp 35 40 45Phe
Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val
Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Arg Val Asn
Asn 85 90 95Lys Ala Leu
Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu
Pro Pro Ser Arg Glu Glu 115 120
125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp Phe Phe 130
135 140Pro Pro Asp Ile Asp Val Glu Trp
Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp
Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210
215 22079220PRTArtificial SequenceSynthetic
Variant canine IgG-C Fc, Protein A -, C1q - , K(93)R 79Pro Gly Cys
Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Val Thr Ala
Arg Thr Pro Thr Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asn Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Ser Lys Gln Val
Gln Thr Ala Asn Thr Gln Pro Arg Glu 50 55
60Glu Gln Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp
Leu Ser Gly Lys Gln Phe Lys Cys Arg Val Asn Asn 85
90 95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile
Ile Ser Lys Thr Pro Gly 100 105
110Gln Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu
115 120 125Met Ser Lys Asn Thr Val Thr
Leu Thr Cys Leu Val Lys Asp Phe Phe 130 135
140Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Ile
Ser Leu Ser His Ser Pro Gly Lys 210 215
22080237PRTUnknownAlternative wild-type feline IgG1a Fc, Protein A
+, C1q + 80Arg Lys Thr Asp His Pro Pro Gly Pro Lys Thr Gly Glu Gly Pro
Lys1 5 10 15Cys Pro Pro
Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro 20
25 30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser
Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50
55 60Trp Phe Val Asp Asn Thr Gln Val Tyr
Thr Ala Lys Thr Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu
Pro Ile 85 90 95Leu His
Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu
Arg Thr Ile Ser Lys Ala Lys 115 120
125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu
130 135 140Glu Leu Ser Glu Asn Lys Val
Ser Val Thr Cys Leu Ile Lys Ser Phe145 150
155 160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr
Gly Gln Pro Glu 165 170
175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly
180 185 190Thr Tyr Phe Val Tyr Ser
Lys Leu Ser Val Asp Arg Ser His Trp Gln 195 200
205Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu
His Ser 210 215 220His His Thr Gln Lys
Ser Leu Thr Gln Ser Pro Gly Lys225 230
23581237PRTUnknownAlternative wild-type feline IgG1b Fc, Protein A
+, C1q + 81Arg Lys Thr Asp His Pro Pro Gly Pro Lys Thr Gly Glu Gly Pro
Lys1 5 10 15Cys Pro Pro
Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro 20
25 30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser
Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50
55 60Trp Phe Val Asp Asn Thr Gln Val Tyr
Thr Ala Lys Thr Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu
Pro Ile 85 90 95Leu His
Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu
Arg Thr Ile Ser Lys Asp Lys 115 120
125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu
130 135 140Glu Leu Ser Glu Asn Lys Val
Ser Val Thr Cys Leu Ile Glu Gly Phe145 150
155 160Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ile Thr
Gly Gln Pro Glu 165 170
175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly
180 185 190Thr Tyr Phe Leu Tyr Ser
Arg Leu Ser Val Asp Arg Ser Arg Trp Gln 195 200
205Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu
His Ser 210 215 220His His Thr Gln Lys
Ser Leu Thr Gln Ser Pro Gly Lys225 230
23582237PRTArtificial SequenceSynthetic Variant feline IgG1a Fc, Protein
A +, C1q -, P(198)A 82Arg Lys Thr Asp His Pro Pro Gly Pro Lys Thr
Gly Glu Gly Pro Lys1 5 10
15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro
20 25 30Pro Lys Pro Lys Asp Thr Leu
Ser Ile Ser Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile
Thr 50 55 60Trp Phe Val Asp Asn Thr
Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65 70
75 80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val
Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn
100 105 110Ser Lys Ser Leu Pro Ser
Pro Ile Glu Arg Thr Ile Ser Lys Ala Lys 115 120
125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala
Gln Glu 130 135 140Glu Leu Ser Glu Asn
Lys Val Ser Val Thr Cys Leu Ile Lys Ser Phe145 150
155 160His Pro Pro Asp Ile Ala Val Glu Trp Glu
Ile Thr Gly Gln Pro Glu 165 170
175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly
180 185 190Thr Tyr Phe Val Tyr
Ser Lys Leu Ser Val Asp Arg Ser His Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu
Ala Leu His Ser 210 215 220His His Thr
Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225 230
23583237PRTArtificial SequenceSynthetic Variant feline IgG1b Fc,
Protein A +, C1q -, P(198)A 83Arg Lys Thr Asp His Pro Pro Gly Pro
Lys Thr Gly Glu Gly Pro Lys1 5 10
15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe
Pro 20 25 30Pro Lys Pro Lys
Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 35
40 45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp
Val Gln Ile Thr 50 55 60Trp Phe Val
Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65 70
75 80Glu Glu Gln Phe Asn Ser Thr Tyr
Arg Val Val Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys
Val Asn 100 105 110Ser Lys Ser
Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Asp Lys 115
120 125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu
Pro Pro Ala Gln Glu 130 135 140Glu Leu
Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Glu Gly Phe145
150 155 160Tyr Pro Ser Asp Ile Ala Val
Glu Trp Glu Ile Thr Gly Gln Pro Glu 165
170 175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly 180 185 190Thr
Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser Arg Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His Ser 210 215
220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225
230 23584220PRTArtificial SequenceSynthetic Variant
canine IgG-C Fc , C1q - , K(93)R , Protein A +, I(21)T, V(23)L,
T(24)I 84Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp
Thr Leu Leu Ile Ala Arg Thr Pro Thr Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu Asn Pro Glu
Val Gln Ile Ser Trp 35 40 45Phe
Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50
55 60Glu Gln Ser Asn Gly Thr Tyr Arg Val Val
Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Arg Val Asn
Asn 85 90 95Lys Ala Leu
Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly 100
105 110Gln Ala His Gln Pro Asn Val Tyr Val Leu
Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe 130
135 140Pro Pro Glu Ile Asp Val Glu Trp
Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp
Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210
215 2208531PRTUnknownWild-type GLP1 (7-37)
85His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1
5 10 15Gln Ala Ala Lys Glu Phe
Ile Ala Trp Leu Val Lys Gly Ala Gly 20 25
308629PRTArtificial SequenceSynthetic GLP1-S8 (7-35) 86His
Ser Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1
5 10 15Gln Ala Ala Lys Glu Phe Ile
Ala Trp Leu Val Lys Gly 20
258729PRTArtificial SequenceSynthetic GLP1-G8 (7-35) 87His Gly Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu
Val Lys Gly 20 25881PRTArtificial
SequenceSynthetic 1G extension 88Gly1892PRTArtificial SequenceSynthetic
2G extension 89Gly Gly1903PRTArtificial SequenceSynthetic 3G extension
90Gly Gly Gly1914PRTArtificial SequenceSynthetic 4G extension 91Gly Gly
Gly Gly1925PRTArtificial SequenceSynthetic 5G extension 92Gly Gly Gly Gly
Gly1 5936PRTArtificial SequenceSynthetic 6G extension 93Gly
Gly Gly Gly Gly Gly1 5947PRTArtificial SequenceSynthetic 7G
extension 94Gly Gly Gly Gly Gly Gly Gly1 5958PRTArtificial
SequenceSynthetic 8G extension 95Gly Gly Gly Gly Gly Gly Gly Gly1
596305PRTArtificial SequenceSynthetic ssGLP1-G8_I_WTfeIgG2 96Met
Ala Val Leu Gly Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1
5 10 15Val Leu Ser His Gly Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val
Lys Gly 35 40 45Gly Ala Gly Gly
Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser 50 55
60Gly Gly Gly Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser
Lys Thr Gly65 70 75
80Glu Gly Pro Lys Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val
85 90 95Phe Ile Phe Pro Pro Lys
Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr 100
105 110Pro Glu Val Thr Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asn 115 120 125Val Gln
Ile Thr Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys 130
135 140Thr Arg Pro Arg Glu Glu Gln Phe Asn Ser Thr
Tyr Arg Val Val Ser145 150 155
160Val Leu Pro Ile Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys
165 170 175Cys Lys Val Asn
Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile 180
185 190Ser Lys Ala Lys Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro 195 200 205Pro
Thr Gln Glu Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu 210
215 220Ile Lys Gly Phe His Pro Pro Asp Ile Ala
Val Glu Trp Glu Ile Thr225 230 235
240Gly Gln Pro Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln
Leu 245 250 255Asp Ser Asp
Gly Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg 260
265 270Ser His Trp Gln Arg Gly Asn Thr Tyr Thr
Cys Ser Val Ser His Glu 275 280
285Ala Leu His Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly 290
295 300Lys30597346PRTArtificial
SequenceSynthetic ssGLP1-G8/GLP1-3G_III_ WTfeIgG2 97Met Ala Val Leu Gly
Leu Leu Phe Cys Leu Val Thr Phe Pro Ser Cys1 5
10 15Val Leu Ser His Gly Glu Gly Thr Phe Thr Ser
Asp Val Ser Ser Tyr 20 25
30Leu Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly
35 40 45Gly Ala Gly Gly Gly Gly Gly Ser
Gly Gly Gly Ser Gly Gly Gly Ser 50 55
60Gly Gly Gly Ser Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly65
70 75 80Glu Gly Pro Lys Cys
Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val 85
90 95Phe Ile Phe Pro Pro Lys Pro Lys Asp Thr Leu
Ser Ile Ser Arg Thr 100 105
110Pro Glu Val Thr Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn
115 120 125Val Gln Ile Thr Trp Phe Val
Asp Asn Thr Glu Met His Thr Ala Lys 130 135
140Thr Arg Pro Arg Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val
Ser145 150 155 160Val Leu
Pro Ile Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys
165 170 175Cys Lys Val Asn Ser Lys Ser
Leu Pro Ser Ala Met Glu Arg Thr Ile 180 185
190Ser Lys Ala Lys Gly Gln Pro His Glu Pro Gln Val Tyr Val
Leu Pro 195 200 205Pro Thr Gln Glu
Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu 210
215 220Ile Lys Gly Phe His Pro Pro Asp Ile Ala Val Glu
Trp Glu Ile Thr225 230 235
240Gly Gln Pro Glu Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu
245 250 255Asp Ser Asp Gly Thr
Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg 260
265 270Ser His Trp Gln Arg Gly Asn Thr Tyr Thr Cys Ser
Val Ser His Glu 275 280 285Ala Leu
His Ser His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly 290
295 300Lys Gly Gly Gly Gly Ser Gly Gly Gly Gly His
Ala Glu Gly Thr Phe305 310 315
320Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly Gln Ala Ala Lys Glu Phe
325 330 335Ile Ala Trp Leu
Val Lys Gly Gly Gly Gly 340
3459830PRTArtificial SequenceSynthetic Variant GLP1 (7-36) 98His Ala Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp
Leu Val Lys Gly Ala 20 25
309929PRTArtificial SequenceSynthetic Variant GLP1 (7-35) 99His Ala Glu
Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp
Leu Val Lys Gly 20
25100229PRTUnknownWild-type equine Fc IgG2 (including equine IgG2
hinge), Protein A - , C1q - 100Val Pro Lys Pro Gln Cys Pro Pro Tyr Thr
His Ser Lys Phe Leu Gly1 5 10
15Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala Leu Met
20 25 30Ile Ser Arg Thr Pro Val
Val Thr Cys Val Val Val Asn Leu Ser Asp 35 40
45Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr
Glu Val 50 55 60His Ser Ala Ile Thr
Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr65 70
75 80Arg Val Val Ser Val Leu Pro Ile Gln His
Gln Asp Trp Leu Ser Gly 85 90
95Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile
100 105 110Ser Arg Ala Ile Ser
Arg Gly Lys Gly Pro Ser Arg Val Pro Gln Val 115
120 125Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys
Ser Lys Val Ser 130 135 140Val Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu145
150 155 160Trp Gln Ser Asn Arg Trp Pro
Glu Leu Glu Gly Lys Tyr Ser Thr Thr 165
170 175Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu
Tyr Ser Lys Leu 180 185 190Ser
Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala 195
200 205Val Met His Glu Ala Leu His Asn His
Phe Thr Lys Thr Asp Ile Ser 210 215
220Glu Ser Leu Gly Lys22510114PRTUnknownWild-type equine IgG1 hinge
101Asp Met Ser Lys Cys Pro Lys Cys Pro Ala Pro Glu Leu Leu1
5 1010215PRTUnknownWild-type equine IgG2 hinge 102Val
Pro Lys Pro Gln Cys Pro Pro Tyr Thr His Ser Lys Phe Leu1 5
10 15103318PRTArtificial
SequenceSynthetic GLP1-G8/GLP1-3G_III_ VAReqIgG2 103His Gly Glu Gly Thr
Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val
Lys Gly Gly Ala Gly 20 25
30Gly Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly
35 40 45Ser Asp Met Ser Lys Cys Pro Lys
Cys Pro Ala Pro Glu Leu Leu Gly 50 55
60Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Thr Leu Met65
70 75 80Ile Ser Arg Thr Pro
Val Val Thr Cys Val Val Val Asn Leu Ser Asp 85
90 95Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val
Asp Asn Thr Glu Val 100 105
110His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr
115 120 125Arg Val Val Ser Val Leu Pro
Ile Gln His Gln Asp Trp Leu Ser Gly 130 135
140Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro
Ile145 150 155 160Ser Arg
Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln Val
165 170 175Tyr Val Leu Pro Pro His Pro
Asp Glu Leu Ala Lys Ser Lys Val Ser 180 185
190Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser
Val Glu 195 200 205Trp Gln Ser Asn
Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr Thr 210
215 220Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu
Tyr Ser Lys Leu225 230 235
240Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala
245 250 255Val Met His Glu Ala
Leu His Asn His Tyr Thr Lys Thr Asp Ile Ser 260
265 270Glu Ser Leu Gly Lys Gly Gly Gly Gly Ser Gly Gly
Gly Gly His Ala 275 280 285Glu Gly
Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly Gln Ala 290
295 300Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly
Gly Gly Gly305 310 315104279PRTArtificial
SequenceSynthetic GLP1-G8_I_VAReqIgG2 104His Gly Glu Gly Thr Phe Thr Ser
Asp Val Ser Ser Tyr Leu Glu Gly1 5 10
15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly
Ala Gly 20 25 30Gly Gly Gly
Gly Ser Gly Gly Gly Ser Gly Gly Gly Ser Gly Gly Gly 35
40 45Ser Ser Val Pro Lys Pro Gln Cys Pro Pro Tyr
Thr His Ser Lys Phe 50 55 60Leu Gly
Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Thr65
70 75 80Leu Met Ile Ser Arg Thr Pro
Val Val Thr Cys Val Val Val Asn Leu 85 90
95Ser Asp Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val
Asp Asn Thr 100 105 110Glu Val
His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser 115
120 125Thr Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Leu 130 135 140Ser
Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln145
150 155 160Pro Ile Ser Arg Ala Ile
Ser Arg Gly Lys Gly Pro Ser Arg Val Pro 165
170 175Gln Val Tyr Val Leu Pro Pro His Pro Asp Glu Leu
Ala Lys Ser Lys 180 185 190Val
Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser 195
200 205Val Glu Trp Gln Ser Asn Arg Trp Pro
Glu Leu Glu Gly Lys Tyr Ser 210 215
220Thr Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser225
230 235 240Lys Leu Ser Leu
Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr 245
250 255Cys Ala Val Met His Glu Ala Leu His Asn
His Tyr Thr Lys Thr Asp 260 265
270Ile Ser Glu Ser Leu Gly Lys 275105321PRTArtificial
SequenceSynthetic GLP1-G8/GLP1-2G_III_ VARcaIgGD 105His Gly Glu Gly Thr
Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu Gly1 5
10 15Gln Ala Ala Lys Glu Phe Ile Ala Trp Leu Val
Lys Gly Gly Gly Gly 20 25
30Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly Ser
35 40 45Pro Lys Glu Ser Thr Cys Lys Cys
Ile Ser Pro Cys Pro Val Pro Glu 50 55
60Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp65
70 75 80Thr Leu Leu Ile Ala
Arg Thr Pro Glu Ile Thr Cys Val Val Leu Asp 85
90 95Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser
Trp Phe Val Asp Gly 100 105
110Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu Gln Gln Phe Asn
115 120 125Ser Thr Tyr Arg Val Val Ser
Val Leu Pro Ile Gly His Gln Asp Trp 130 135
140Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His Ile Gly Leu
Pro145 150 155 160Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly Gln Ala His Gln
165 170 175Pro Ser Val Tyr Val Leu Pro
Pro Ser Pro Lys Glu Leu Ser Ser Ser 180 185
190Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe Phe Pro
Pro Glu 195 200 205Ile Asp Val Glu
Trp Gln Ser Asn Gly Gln Pro Glu Pro Glu Ser Lys 210
215 220Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
Ser Tyr Phe Leu225 230 235
240Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Gln Gly Asp Thr
245 250 255Phe Thr Cys Ala Val
Met His Glu Ala Leu His Asn His Tyr Thr Asp 260
265 270Leu Ser Leu Ser His Ser Pro Gly Lys Gly Gly Gly
Gly Ser Gly Gly 275 280 285Gly Gly
His Ala Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu 290
295 300Glu Gly Gln Ala Ala Lys Glu Phe Ile Ala Trp
Leu Val Lys Gly Gly305 310 315
320Gly106321PRTArtificial SequenceSynthetic GLP1-G8/GLP1-2G_III_
VARcaIgGD 106His Gly Glu Gly Thr Phe Thr Ser Asp Val Ser Ser Tyr Leu Glu
Gly1 5 10 15Gln Ala Ala
Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly Gly Gly 20
25 30Ser Gly Gly Gly Gly Ser Gly Gly Gly Gly
Ser Gly Gly Gly Gly Ser 35 40
45Pro Lys Glu Ser Thr Ser Lys Cys Ile Ser Pro Cys Pro Val Pro Glu 50
55 60Ser Leu Gly Gly Pro Ser Val Phe Ile
Phe Pro Pro Lys Pro Lys Asp65 70 75
80Thr Leu Leu Ile Ala Arg Thr Pro Glu Ile Thr Cys Val Val
Leu Asp 85 90 95Leu Gly
Arg Glu Asp Pro Glu Val Gln Ile Ser Trp Phe Val Asp Gly 100
105 110Lys Glu Val His Thr Ala Lys Thr Gln
Pro Arg Glu Gln Gln Phe Asn 115 120
125Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly His Gln Asp Trp
130 135 140Leu Thr Gly Lys Glu Phe Lys
Cys Arg Val Asn His Ile Gly Leu Pro145 150
155 160Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
Gln Ala His Gln 165 170
175Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu Leu Ser Ser Ser
180 185 190Asp Thr Val Thr Leu Thr
Cys Leu Ile Lys Asp Phe Phe Pro Pro Glu 195 200
205Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro Glu Pro Glu
Ser Lys 210 215 220Tyr His Thr Thr Ala
Pro Gln Leu Asp Glu Asp Gly Ser Tyr Phe Leu225 230
235 240Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg
Trp Gln Gln Gly Asp Thr 245 250
255Phe Thr Cys Ala Val Met His Glu Ala Leu His Asn His Tyr Thr Asp
260 265 270Leu Ser Leu Ser His
Ser Pro Gly Lys Gly Gly Gly Gly Ser Gly Gly 275
280 285Gly Gly His Ala Glu Gly Thr Phe Thr Ser Asp Val
Ser Ser Tyr Leu 290 295 300Glu Gly Gln
Ala Ala Lys Glu Phe Ile Ala Trp Leu Val Lys Gly Gly305
310 315 320Gly107237PRTUnknownWild-type
canine IgG-B Fc with hinge, Protein A +, C1q +, CD16 + 107Pro Lys
Arg Glu Asn Gly Arg Val Pro Arg Pro Pro Asp Cys Pro Lys1 5
10 15Cys Pro Ala Pro Glu Met Leu Gly
Gly Pro Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val
Thr 35 40 45Cys Val Val Val Asp
Leu Asp Pro Glu Asp Pro Glu Val Gln Ile Ser 50 55
60Trp Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln
Pro Arg65 70 75 80Glu
Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Gly His Gln Asp Trp Leu Lys
Gly Lys Gln Phe Thr Cys Lys Val Asn 100 105
110Asn Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys
Ala Arg 115 120 125Gly Gln Ala His
Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu 130
135 140Glu Leu Ser Lys Asn Thr Val Ser Leu Thr Cys Leu
Ile Lys Asp Phe145 150 155
160Phe Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
165 170 175Pro Glu Ser Lys Tyr
Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly 180
185 190Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys
Ser Arg Trp Gln 195 200 205Arg Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn 210
215 220His Tyr Thr Gln Glu Ser Leu Ser His Ser Pro
Gly Lys225 230
235108235PRTUnknownWild-type canine IgG-C Fc with hinge, Protein A ,
C1q +, CD16 + 108Ala Lys Glu Cys Glu Cys Lys Cys Asn Cys Asn Asn Cys Pro
Cys Pro1 5 10 15Gly Cys
Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys 20
25 30Pro Lys Asp Ile Leu Val Thr Ala Arg
Thr Pro Thr Val Thr Cys Val 35 40
45Val Val Asp Leu Asp Pro Glu Asn Pro Glu Val Gln Ile Ser Trp Phe 50
55 60Val Asp Ser Lys Gln Val Gln Thr Ala
Asn Thr Gln Pro Arg Glu Glu65 70 75
80Gln Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile
Gly His 85 90 95Gln Asp
Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val Asn Asn Lys 100
105 110Ala Leu Pro Ser Pro Ile Glu Glu Ile
Ile Ser Lys Thr Pro Gly Gln 115 120
125Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu Met
130 135 140Ser Lys Asn Thr Val Thr Leu
Thr Cys Leu Val Lys Asp Phe Phe Pro145 150
155 160Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln
Gln Glu Pro Glu 165 170
175Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser Tyr
180 185 190Phe Leu Tyr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg Gly 195 200
205Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn
His Tyr 210 215 220Thr Gln Ile Ser Leu
Ser His Ser Pro Gly Lys225 230
235109221PRTArtificial SequenceVariant canine IgG-A Fc, Heterodimer chain
3, T(138)W 109Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val Leu Ile
Phe Pro Pro1 5 10 15Lys
Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu
Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg Glu
50 55 60Gln Gln Phe Asn Gly Thr Tyr Arg
Val Val Ser Val Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg
Val Asn His 85 90 95Ile
Asp Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Arg Ala His Lys Pro Ser Val
Tyr Val Leu Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Ser Ile Trp Cys Leu Ile Lys Asp
Phe 130 135 140Tyr Pro Pro Asp Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu145 150
155 160Pro Glu Arg Lys His Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Pro Phe Thr
Cys Ala Val Met His Glu Thr Leu Gln Asn 195 200
205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
210 215 220110221PRTArtificial
SequenceVariant canine IgG-A Fc, Heterodimer chain 4, T(138)S,
L(140)A, Y(181)T 110Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val Leu Ile
Phe Pro Pro1 5 10 15Lys
Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu
Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg Glu
50 55 60Gln Gln Phe Asn Gly Thr Tyr Arg
Val Val Ser Val Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg
Val Asn His 85 90 95Ile
Asp Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Arg Ala His Lys Pro Ser Val
Tyr Val Leu Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Ser Ile Ser Cys Ala Ile Lys Asp
Phe 130 135 140Tyr Pro Pro Asp Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu145 150
155 160Pro Glu Arg Lys His Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Thr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Pro Phe Thr
Cys Ala Val Met His Glu Thr Leu Gln Asn 195 200
205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
210 215 220111220PRTArtificial
SequenceVariant canine IgG-B Fc, Heterodimer chain 3, T(137)W 111Pro
Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Thr Leu Leu
Ile Ala Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln Ile
Ser Trp 35 40 45Phe Val Asp Gly
Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu
Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn
85 90 95Lys Ala Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro
Ser Arg Glu Glu 115 120 125Leu Ser
Lys Asn Thr Val Ser Leu Trp Cys Leu Ile Lys Asp Phe Phe 130
135 140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210 215
220112220PRTArtificial SequenceVariant canine IgG-B Fc,
Heterodimer chain 4, T(137)S, L(139)A, Y(180)T 112Pro Ala Pro Glu
Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg
Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Gln Met Gln
Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu
Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn 85
90 95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu
115 120 125Leu Ser Lys Asn Thr Val Ser
Leu Ser Cys Ala Ile Lys Asp Phe Phe 130 135
140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Thr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Glu
Ser Leu Ser His Ser Pro Gly Lys 210 215
220113220PRTArtificial SequenceVariant canine IgG-C Fc, Heterodimer
chain 3, T(137)W 113Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg
Glu 50 55 60Glu Gln Ser Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys
Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro Asn
Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Trp Cys Leu Val Lys Asp
Phe Phe 130 135 140Pro Pro Glu Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys
210 215 220114220PRTArtificial
SequenceVariant canine IgG-C Fc, Heterodimer chain 4, T(137)S,
L(139)A, Y(180)T 114Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile
Phe Pro Pro1 5 10 15Lys
Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu
Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu
50 55 60Glu Gln Ser Asn Gly Thr Tyr Arg
Val Val Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys
Val Asn Asn 85 90 95Lys
Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro Asn Val
Tyr Val Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Ser Cys Ala Val Lys Asp Phe
Phe 130 135 140Pro Pro Glu Ile Asp Val
Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu
Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Thr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210
215 220115221PRTArtificial SequenceVariant
canine IgG-D Fc, Heterodimer chain 3, T(138)W 115Pro Val Pro Glu Ser
Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr
Pro Glu Ile Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Glu Val His
Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Gln Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65
70 75 80His Gln Asp Trp Leu
Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu
115 120 125Leu Ser Ser Ser Asp Thr Val
Thr Leu Trp Cys Leu Ile Lys Asp Phe 130 135
140Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro
Glu145 150 155 160Pro Glu
Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln 180 185
190Gln Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu
Gln Asn 195 200 205His Tyr Thr Asp
Leu Ser Leu Ser His Ser Pro Gly Lys 210 215
220116221PRTArtificial SequenceVariant canine IgG-D Fc, Heterodimer
chain 4, T(138)S, L(140)A, Y(181)T 116Pro Val Pro Glu Ser Leu Gly
Gly Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu
Ile Thr Cys 20 25 30Val Val
Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Gly Lys Glu Val His Thr Ala
Lys Thr Gln Pro Arg Glu 50 55 60Gln
Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65
70 75 80His Gln Asp Trp Leu Thr
Gly Lys Glu Phe Lys Cys Arg Val Asn His 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser
Lys Ala Arg Gly 100 105 110Gln
Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu 115
120 125Leu Ser Ser Ser Asp Thr Val Thr Leu
Ser Cys Ala Ile Lys Asp Phe 130 135
140Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro Glu145
150 155 160Pro Glu Ser Lys
Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly 165
170 175Ser Tyr Phe Leu Thr Ser Lys Leu Ser Val
Asp Lys Ser Arg Trp Gln 180 185
190Gln Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu Gln Asn
195 200 205His Tyr Thr Asp Leu Ser Leu
Ser His Ser Pro Gly Lys 210 215
220117237PRTUnknownWild-type feline IgG1a Fc, Protein A +, C1q + 117Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Ser Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Val Tyr Ser Lys Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230
235118237PRTUnknownWild-type feline IgG1b Fc, Protein A +, C1q + 118Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Asp Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Glu Gly Phe145 150 155
160Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser Arg Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235119237PRTArtificial
SequenceVariant feline IgG2 Fc, Heterodimer chain 1, T(154)W 119Pro
Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Lys1
5 10 15Cys Pro Val Pro Glu Ile Pro
Gly Ala Pro Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr
Arg Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Trp Cys
Leu Ile Lys Gly Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235120237PRTArtificial
SequenceVariant feline IgG2 Fc, Heterodimer chain 2, T(154)S,
L(156)A, Y(197)T 120Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu
Gly Pro Lys1 5 10 15Cys
Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe Ile Phe Pro 20
25 30Pro Lys Pro Lys Asp Thr Leu Ser
Ile Ser Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr
50 55 60Trp Phe Val Asp Asn Thr Glu Met
His Thr Ala Lys Thr Arg Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val
Leu Pro Ile 85 90 95Leu
His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn
100 105 110Ser Lys Ser Leu Pro Ser Ala
Met Glu Arg Thr Ile Ser Lys Ala Lys 115 120
125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln
Glu 130 135 140Glu Leu Ser Glu Asn Lys
Val Ser Val Ser Cys Ala Ile Lys Gly Phe145 150
155 160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile
Thr Gly Gln Pro Glu 165 170
175Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly
180 185 190Thr Tyr Phe Leu Thr Ser
Arg Leu Ser Val Asp Arg Ser His Trp Gln 195 200
205Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu
His Ser 210 215 220His His Thr Gln Lys
Ser Leu Thr Gln Ser Pro Gly Lys225 230
235121237PRTArtificial SequenceVariant feline IgG1a Fc, Heterodimer chain
1, T(154)W 121Arg Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp
Cys Pro Lys1 5 10 15Cys
Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro 20
25 30Pro Lys Pro Lys Asp Thr Leu Ser
Ile Ser Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr
50 55 60Trp Phe Val Asp Asn Thr Gln Val
Tyr Thr Ala Lys Thr Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val
Leu Pro Ile 85 90 95Leu
His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn
100 105 110Ser Lys Ser Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Lys 115 120
125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln
Glu 130 135 140Glu Leu Ser Glu Asn Lys
Val Ser Val Trp Cys Leu Ile Lys Ser Phe145 150
155 160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile
Thr Gly Gln Pro Glu 165 170
175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly
180 185 190Thr Tyr Phe Val Tyr Ser
Lys Leu Ser Val Asp Arg Ser His Trp Gln 195 200
205Arg Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu
His Ser 210 215 220His His Thr Gln Lys
Ser Leu Thr Gln Ser Pro Gly Lys225 230
235122237PRTArtificial SequenceVariant feline IgG1a Fc, Heterodimer chain
2, T(154)S, L(156)A, Y(197)T 122Arg Lys Thr Asp His Pro Pro Gly Pro
Lys Pro Cys Asp Cys Pro Lys1 5 10
15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe
Pro 20 25 30Pro Lys Pro Lys
Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 35
40 45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp
Val Gln Ile Thr 50 55 60Trp Phe Val
Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65 70
75 80Glu Glu Gln Phe Asn Ser Thr Tyr
Arg Val Val Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys
Val Asn 100 105 110Ser Lys Ser
Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Lys 115
120 125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu
Pro Pro Ala Gln Glu 130 135 140Glu Leu
Ser Glu Asn Lys Val Ser Val Ser Cys Ala Ile Lys Ser Phe145
150 155 160His Pro Pro Asp Ile Ala Val
Glu Trp Glu Ile Thr Gly Gln Pro Glu 165
170 175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly 180 185 190Thr
Tyr Phe Val Thr Ser Lys Leu Ser Val Asp Arg Ser His Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His Ser 210 215
220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225
230 235123237PRTArtificial SequenceVariant feline IgG1b
Fc, Heterodimer chain 1, T(154)W 123Arg Lys Thr Asp His Pro Pro Gly
Pro Lys Pro Cys Asp Cys Pro Lys1 5 10
15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile
Phe Pro 20 25 30Pro Lys Pro
Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 35
40 45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser
Asp Val Gln Ile Thr 50 55 60Trp Phe
Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65
70 75 80Glu Glu Gln Phe Asn Ser Thr
Tyr Arg Val Val Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys
Lys Val Asn 100 105 110Ser Lys
Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Asp Lys 115
120 125Gly Gln Pro His Glu Pro Gln Val Tyr Val
Leu Pro Pro Ala Gln Glu 130 135 140Glu
Leu Ser Glu Asn Lys Val Ser Val Trp Cys Leu Ile Glu Gly Phe145
150 155 160Tyr Pro Ser Asp Ile Ala
Val Glu Trp Glu Ile Thr Gly Gln Pro Glu 165
170 175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly 180 185 190Thr
Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser Arg Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His Ser 210 215
220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225
230 235124237PRTArtificial SequenceVariant feline IgG1b
Fc, Heterodimer chain 2, T(154)S, L(156)A, Y(197)T 124Arg Lys Thr
Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1 5
10 15Cys Pro Pro Pro Glu Met Leu Gly Gly
Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
35 40 45Cys Leu Val Val Asp Leu Gly
Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65
70 75 80Glu Glu Gln Phe
Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile 85
90 95Leu His Gln Asp Trp Leu Lys Gly Lys Glu
Phe Lys Cys Lys Val Asn 100 105
110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Asp Lys
115 120 125Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro Pro Ala Gln Glu 130 135
140Glu Leu Ser Glu Asn Lys Val Ser Val Ser Cys Ala Ile Glu Gly
Phe145 150 155 160Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn Tyr Arg Thr
Thr Pro Pro Gln Leu Asp Ser Asp Gly 180 185
190Thr Tyr Phe Leu Thr Ser Arg Leu Ser Val Asp Arg Ser Arg
Trp Gln 195 200 205Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly
Lys225 230 235125237PRTArtificial
SequenceVariant feline IgG2 Fc, with feline IgG1 hinge 125Arg Lys Thr Asp
His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1 5
10 15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro
Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
35 40 45Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asn Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg Pro Arg65
70 75 80Glu Glu Gln Phe Asn
Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile 85
90 95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe
Lys Cys Lys Val Asn 100 105
110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys Ala Lys
115 120 125Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro Pro Thr Gln Glu 130 135
140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Gly
Phe145 150 155 160His Pro
Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn Tyr Gln Thr
Thr Pro Pro Gln Leu Asp Ser Asp Gly 180 185
190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His
Trp Gln 195 200 205Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly
Lys225 230 235126237PRTArtificial
SequenceVariant feline IgG2 Fc with modified hinge, K(16)P 126Pro
Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Pro1
5 10 15Cys Pro Val Pro Glu Ile Pro
Gly Ala Pro Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr
Arg Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Gly Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235127237PRTArtificial
SequenceVariant feline IgG1a Fc with modified hinge, K(16)P 127Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Pro1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Ser Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Val Tyr Ser Lys Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235128237PRTArtificial
SequenceVariant feline IgG1b Fc with modified hinge, K(16)P 128Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Pro1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Asp Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Glu Gly Phe145 150 155
160Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser Arg Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230
235129244PRTUnknownWild-type equine IgG2 Fc with hinge, Protein A ,
C1q 129Pro Pro Cys Val Leu Ser Ala Glu Gly Val Ile Pro Ile Pro Ser Val1
5 10 15Pro Lys Pro Gln Cys
Pro Pro Tyr Thr His Ser Lys Phe Leu Gly Gly 20
25 30Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp
Ala Leu Met Ile 35 40 45Ser Arg
Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser Asp Gln 50
55 60Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp
Asn Thr Glu Val His65 70 75
80Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr Arg
85 90 95Val Val Ser Val Leu
Pro Ile Gln His Gln Asp Trp Leu Ser Gly Lys 100
105 110Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro
Gln Pro Ile Ser 115 120 125Arg Ala
Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln Val Tyr 130
135 140Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys
Ser Lys Val Ser Val145 150 155
160Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp
165 170 175Gln Ser Asn Arg
Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr Thr Pro 180
185 190Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu
Tyr Ser Lys Leu Ser 195 200 205Leu
Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala Val 210
215 220Met His Glu Ala Leu His Asn His Phe Thr
Lys Thr Asp Ile Ser Glu225 230 235
240Ser Leu Gly Lys130244PRTArtificial SequenceVariant equine
IgG2 Fc with modified hinge, Protein A , C1q , Q(20)P 130Pro Pro Cys
Val Leu Ser Ala Glu Gly Val Ile Pro Ile Pro Ser Val1 5
10 15Pro Lys Pro Pro Cys Pro Pro Tyr Thr
His Ser Lys Phe Leu Gly Gly 20 25
30Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala Leu Met Ile
35 40 45Ser Arg Thr Pro Val Val Thr
Cys Val Val Val Asn Leu Ser Asp Gln 50 55
60Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val His65
70 75 80Ser Ala Ile Thr
Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr Arg 85
90 95Val Val Ser Val Leu Pro Ile Gln His Gln
Asp Trp Leu Ser Gly Lys 100 105
110Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile Ser
115 120 125Arg Ala Ile Ser Arg Gly Lys
Gly Pro Ser Arg Val Pro Gln Val Tyr 130 135
140Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
Val145 150 155 160Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp
165 170 175Gln Ser Asn Arg Trp Pro Glu
Leu Glu Gly Lys Tyr Ser Thr Thr Pro 180 185
190Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser 195 200 205Leu Glu Thr Ser
Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala Val 210
215 220Met His Glu Ala Leu His Asn His Phe Thr Lys Thr
Asp Ile Ser Glu225 230 235
240Ser Leu Gly Lys131244PRTArtificial SequenceVariant equine IgG2 Fc
with modified hinge, Protein A , C1q , C(3)S 131Pro Pro Ser Val Leu
Ser Ala Glu Gly Val Ile Pro Ile Pro Ser Val1 5
10 15Pro Lys Pro Gln Cys Pro Pro Tyr Thr His Ser
Lys Phe Leu Gly Gly 20 25
30Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala Leu Met Ile
35 40 45Ser Arg Thr Pro Val Val Thr Cys
Val Val Val Asn Leu Ser Asp Gln 50 55
60Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val His65
70 75 80Ser Ala Ile Thr Lys
Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr Arg 85
90 95Val Val Ser Val Leu Pro Ile Gln His Gln Asp
Trp Leu Ser Gly Lys 100 105
110Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile Ser
115 120 125Arg Ala Ile Ser Arg Gly Lys
Gly Pro Ser Arg Val Pro Gln Val Tyr 130 135
140Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
Val145 150 155 160Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp
165 170 175Gln Ser Asn Arg Trp Pro Glu
Leu Glu Gly Lys Tyr Ser Thr Thr Pro 180 185
190Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser 195 200 205Leu Glu Thr Ser
Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala Val 210
215 220Met His Glu Ala Leu His Asn His Phe Thr Lys Thr
Asp Ile Ser Glu225 230 235
240Ser Leu Gly Lys132244PRTArtificial SequenceVariant equine IgG2 Fc
with modified hinge, Protein A , C1q , C(3)S, Q(20)P 132Pro Pro Ser
Val Leu Ser Ala Glu Gly Val Ile Pro Ile Pro Ser Val1 5
10 15Pro Lys Pro Pro Cys Pro Pro Tyr Thr
His Ser Lys Phe Leu Gly Gly 20 25
30Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala Leu Met Ile
35 40 45Ser Arg Thr Pro Val Val Thr
Cys Val Val Val Asn Leu Ser Asp Gln 50 55
60Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val His65
70 75 80Ser Ala Ile Thr
Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr Arg 85
90 95Val Val Ser Val Leu Pro Ile Gln His Gln
Asp Trp Leu Ser Gly Lys 100 105
110Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile Ser
115 120 125Arg Ala Ile Ser Arg Gly Lys
Gly Pro Ser Arg Val Pro Gln Val Tyr 130 135
140Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
Val145 150 155 160Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp
165 170 175Gln Ser Asn Arg Trp Pro Glu
Leu Glu Gly Lys Tyr Ser Thr Thr Pro 180 185
190Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser 195 200 205Leu Glu Thr Ser
Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala Val 210
215 220Met His Glu Ala Leu His Asn His Phe Thr Lys Thr
Asp Ile Ser Glu225 230 235
240Ser Leu Gly Lys133244PRTArtificial SequenceVariant equine IgG2 Fc
with hinge, Protein A +, C1q , A(45)T, F(233)Y 133Pro Pro Cys Val
Leu Ser Ala Glu Gly Val Ile Pro Ile Pro Ser Val1 5
10 15Pro Lys Pro Gln Cys Pro Pro Tyr Thr His
Ser Lys Phe Leu Gly Gly 20 25
30Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Thr Leu Met Ile
35 40 45Ser Arg Thr Pro Val Val Thr Cys
Val Val Val Asn Leu Ser Asp Gln 50 55
60Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu Val His65
70 75 80Ser Ala Ile Thr Lys
Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr Arg 85
90 95Val Val Ser Val Leu Pro Ile Gln His Gln Asp
Trp Leu Ser Gly Lys 100 105
110Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro Ile Ser
115 120 125Arg Ala Ile Ser Arg Gly Lys
Gly Pro Ser Arg Val Pro Gln Val Tyr 130 135
140Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val Ser
Val145 150 155 160Thr Cys
Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp
165 170 175Gln Ser Asn Arg Trp Pro Glu
Leu Glu Gly Lys Tyr Ser Thr Thr Pro 180 185
190Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser 195 200 205Leu Glu Thr Ser
Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala Val 210
215 220Met His Glu Ala Leu His Asn His Tyr Thr Lys Thr
Asp Ile Ser Glu225 230 235
240Ser Leu Gly Lys134244PRTArtificial SequenceVariant equine IgG2 Fc
with modified hinge, Protein A +, C1q , Q(20)P, A(45)T, F(233)Y
134Pro Pro Cys Val Leu Ser Ala Glu Gly Val Ile Pro Ile Pro Ser Val1
5 10 15Pro Lys Pro Pro Cys Pro
Pro Tyr Thr His Ser Lys Phe Leu Gly Gly 20 25
30Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Thr
Leu Met Ile 35 40 45Ser Arg Thr
Pro Val Val Thr Cys Val Val Val Asn Leu Ser Asp Gln 50
55 60Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn
Thr Glu Val His65 70 75
80Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser Thr Tyr Arg
85 90 95Val Val Ser Val Leu Pro
Ile Gln His Gln Asp Trp Leu Ser Gly Lys 100
105 110Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro
Gln Pro Ile Ser 115 120 125Arg Ala
Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln Val Tyr 130
135 140Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys
Ser Lys Val Ser Val145 150 155
160Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val Glu Trp
165 170 175Gln Ser Asn Arg
Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr Thr Pro 180
185 190Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu
Tyr Ser Lys Leu Ser 195 200 205Leu
Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys Ala Val 210
215 220Met His Glu Ala Leu His Asn His Tyr Thr
Lys Thr Asp Ile Ser Glu225 230 235
240Ser Leu Gly Lys135244PRTArtificial SequenceVariant equine
IgG2 Fc with modified hinge, Protein A +, C1q , C(3)S, Q(20)P,
A(45)T, F(233)Y 135Pro Pro Ser Val Leu Ser Ala Glu Gly Val Ile Pro Ile
Pro Ser Val1 5 10 15Pro
Lys Pro Pro Cys Pro Pro Tyr Thr His Ser Lys Phe Leu Gly Gly 20
25 30Pro Ser Val Phe Ile Phe Pro Pro
Asn Pro Lys Asp Thr Leu Met Ile 35 40
45Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser Asp Gln
50 55 60Tyr Pro Asp Val Gln Phe Ser Trp
Tyr Val Asp Asn Thr Glu Val His65 70 75
80Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Ser
Thr Tyr Arg 85 90 95Val
Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser Gly Lys
100 105 110Glu Phe Lys Cys Ser Val Thr
Asn Val Gly Val Pro Gln Pro Ile Ser 115 120
125Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln Val
Tyr 130 135 140Val Leu Pro Pro His Pro
Asp Glu Leu Ala Lys Ser Lys Val Ser Val145 150
155 160Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp
Ile Ser Val Glu Trp 165 170
175Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr Thr Pro
180 185 190Ala Gln Leu Asp Gly Asp
Gly Ser Tyr Phe Leu Tyr Ser Lys Leu Ser 195 200
205Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys
Ala Val 210 215 220Met His Glu Ala Leu
His Asn His Tyr Thr Lys Thr Asp Ile Ser Glu225 230
235 240Ser Leu Gly Lys136237PRTArtificial
SequenceVariant feline IgG1a Fc, Protein A +, C1q , P(198)A 136Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Ser Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Val Tyr Ser Lys Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235137237PRTArtificial
SequenceVariant feline IgG1b Fc, Protein A +, C1q , P(198)A 137Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Asn Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Asp Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Glu Gly Phe145 150 155
160Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser Arg Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230
23513830PRTUnknownWild-type equine IgG2 hinge 138Pro Pro Cys Val Leu Ser
Ala Glu Gly Val Ile Pro Ile Pro Ser Val1 5
10 15Pro Lys Pro Gln Cys Pro Pro Tyr Thr His Ser Lys
Phe Leu 20 25
30139220PRTArtificial SequenceVariant canine IgG-B Fc, Protein A +, C1q
+, CD16 , M(5)P 139Pro Ala Pro Glu Pro Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg
Glu 50 55 60Glu Gln Phe Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr
Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Gln Ala His Gln Pro Ser
Val Tyr Val Leu Pro Pro Ser Arg Glu Glu 115 120
125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp
Phe Phe 130 135 140Pro Pro Asp Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys
210 215 220140220PRTArtificial
SequenceVariant canine IgG-B Fc, Protein A +, C1q +, CD16 , P(39)R
140Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Thr Leu
Leu Ile Ala Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Arg Glu Asp Pro Glu Val Gln
Ile Ser Trp 35 40 45Phe Val Asp
Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val
Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn
85 90 95Lys Ala Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro
Ser Arg Glu Glu 115 120 125Leu Ser
Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp Phe Phe 130
135 140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210 215
220141220PRTArtificial SequenceVariant canine IgG-B Fc,
Protein A +, C1q +, CD16 , D(38)G 141Pro Ala Pro Glu Met Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val
Thr Cys 20 25 30Val Val Val
Asp Leu Gly Pro Glu Asp Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys
Thr Gln Pro Arg Glu 50 55 60Glu Gln
Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Lys Gly
Lys Gln Phe Thr Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys
Ala Arg Gly 100 105 110Gln Ala
His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu 115
120 125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys
Leu Ile Lys Asp Phe Phe 130 135 140Pro
Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Thr
Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Glu Ser Leu Ser His Ser
Pro Gly Lys 210 215
220142220PRTArtificial SequenceVariant canine IgG-B Fc, Protein A +, C1q
+, CD16 , K(97)I 142Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg
Glu 50 55 60Glu Gln Phe Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr
Cys Lys Val Asn Asn 85 90
95Ile Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Gln Ala His Gln Pro Ser
Val Tyr Val Leu Pro Pro Ser Arg Glu Glu 115 120
125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp
Phe Phe 130 135 140Pro Pro Asp Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys
210 215 220143220PRTArtificial
SequenceVariant canine IgG-B Fc, Protein A +, C1q +, CD16 , A(98)G
143Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Thr Leu
Leu Ile Ala Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln
Ile Ser Trp 35 40 45Phe Val Asp
Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val
Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn
85 90 95Lys Gly Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro
Ser Arg Glu Glu 115 120 125Leu Ser
Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp Phe Phe 130
135 140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210 215
220144220PRTArtificial SequenceVariant canine IgG-B Fc,
Protein A +, C1q +, CD16 , D(38)G, K(97)I, A(98)G 144Pro Ala Pro Glu
Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg
Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Gly Pro Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Gln Met Gln
Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu
Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu
115 120 125Leu Ser Lys Asn Thr Val Ser
Leu Thr Cys Leu Ile Lys Asp Phe Phe 130 135
140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Glu
Ser Leu Ser His Ser Pro Gly Lys 210 215
220145220PRTArtificial SequenceVariant canine IgG-B Fc, Protein A +, C1q
, CD16 , D(38)G, K(93)R, K(97)I, A(98)G 145Pro Ala Pro Glu Met Leu
Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro
Glu Val Thr Cys 20 25 30Val
Val Val Asp Leu Gly Pro Glu Asp Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Gly Lys Gln Met Gln Thr
Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu
Lys Gly Lys Gln Phe Thr Cys Arg Val Asn Asn 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu
115 120 125Leu Ser Lys Asn Thr Val Ser
Leu Thr Cys Leu Ile Lys Asp Phe Phe 130 135
140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Glu
Ser Leu Ser His Ser Pro Gly Lys 210 215
220146220PRTArtificial SequenceVariant canine IgG-B Fc, Protein A +, C1q
+, CD16 , M(5)P, P(39)R 146Pro Ala Pro Glu Pro Leu Gly Gly Pro Ser
Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val Thr Cys
20 25 30Val Val Val Asp Leu Asp
Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro
Arg Glu 50 55 60Glu Gln Phe Asn Gly
Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Lys Gly Lys Gln Phe
Thr Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Gln Ala His Gln Pro
Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu 115
120 125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys Leu Ile
Lys Asp Phe Phe 130 135 140Pro Pro Asp
Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Thr Thr
Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Glu Ser Leu Ser His Ser
Pro Gly Lys 210 215
220147220PRTArtificial SequenceVariant canine IgG-B Fc, Protein A +, C1q
, CD16 , M(5)P, P(39)R, K(93)R 147Pro Ala Pro Glu Pro Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Thr Leu Leu Ile Ala Arg Thr Pro Glu Val
Thr Cys 20 25 30Val Val Val
Asp Leu Asp Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Gly Lys Gln Met Gln Thr Ala Lys
Thr Gln Pro Arg Glu 50 55 60Glu Gln
Phe Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Lys Gly
Lys Gln Phe Thr Cys Arg Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys
Ala Arg Gly 100 105 110Gln Ala
His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu 115
120 125Leu Ser Lys Asn Thr Val Ser Leu Thr Cys
Leu Ile Lys Asp Phe Phe 130 135 140Pro
Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Thr
Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Glu Ser Leu Ser His Ser
Pro Gly Lys 210 215
220148220PRTArtificial SequenceVariant canine IgG-C Fc, Protein A +, C1q
+, CD16 , L(5)P 148Pro Gly Cys Gly Pro Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg
Glu 50 55 60Glu Gln Ser Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys
Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro Asn
Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp
Phe Phe 130 135 140Pro Pro Glu Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys
210 215 220149220PRTArtificial
SequenceVariant canine IgG-C Fc, Protein A +, C1q +, CD16 , P(39)R
149Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Ile Leu
Val Thr Ala Arg Thr Pro Thr Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Arg Glu Asn Pro Glu Val Gln
Ile Ser Trp 35 40 45Phe Val Asp
Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50
55 60Glu Gln Ser Asn Gly Thr Tyr Arg Val Val Ser Val
Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val Asn Asn
85 90 95Lys Ala Leu Pro Ser Pro
Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly 100
105 110Gln Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro
Ser Arg Asp Glu 115 120 125Met Ser
Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe 130
135 140Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210 215
220150220PRTArtificial SequenceVariant canine IgG-C Fc,
Protein A +, C1q +, CD16 , D(38)G 150Pro Gly Cys Gly Leu Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val
Thr Cys 20 25 30Val Val Val
Asp Leu Gly Pro Glu Asn Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn
Thr Gln Pro Arg Glu 50 55 60Glu Gln
Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Ser Gly
Lys Gln Phe Lys Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys
Thr Pro Gly 100 105 110Gln Ala
His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr Cys
Leu Val Lys Asp Phe Phe 130 135 140Pro
Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Met
Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser
Pro Gly Lys 210 215
220151220PRTArtificial SequenceVariant canine IgG-C Fc, Protein A +, C1q
+, CD16 , K(97)I 151Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe
Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp Pro
Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg
Glu 50 55 60Glu Gln Ser Asn Gly Thr
Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys
Cys Lys Val Asn Asn 85 90
95Ile Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro Asn
Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp
Phe Phe 130 135 140Pro Pro Glu Ile Asp
Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln
Leu Asp Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile
Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys
210 215 220152220PRTArtificial
SequenceVariant canine IgG-C Fc, Protein A +, C1q +, CD16 , A(98)G
152Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Ile Leu
Val Thr Ala Arg Thr Pro Thr Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asn Pro Glu Val Gln
Ile Ser Trp 35 40 45Phe Val Asp
Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50
55 60Glu Gln Ser Asn Gly Thr Tyr Arg Val Val Ser Val
Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val Asn Asn
85 90 95Lys Gly Leu Pro Ser Pro
Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly 100
105 110Gln Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro
Ser Arg Asp Glu 115 120 125Met Ser
Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe 130
135 140Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210 215
220153220PRTArtificial SequenceVariant canine IgG-C Fc,
Protein A +, C1q , CD16 +, K(93)R 153Pro Gly Cys Gly Leu Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val
Thr Cys 20 25 30Val Val Val
Asp Leu Asp Pro Glu Asn Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn
Thr Gln Pro Arg Glu 50 55 60Glu Gln
Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Ser Gly
Lys Gln Phe Lys Cys Arg Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys
Thr Pro Gly 100 105 110Gln Ala
His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr Cys
Leu Val Lys Asp Phe Phe 130 135 140Pro
Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Met
Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser
Pro Gly Lys 210 215
220154220PRTArtificial SequenceVariant canine IgG-C Fc, Protein A +, C1q
+, CD16 , D(38)G, K(97)I, A(98)G 154Pro Gly Cys Gly Leu Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val
Thr Cys 20 25 30Val Val Val
Asp Leu Gly Pro Glu Asn Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn
Thr Gln Pro Arg Glu 50 55 60Glu Gln
Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Ser Gly
Lys Gln Phe Lys Cys Lys Val Asn Asn 85 90
95Ile Gly Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys
Thr Pro Gly 100 105 110Gln Ala
His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr Cys
Leu Val Lys Asp Phe Phe 130 135 140Pro
Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Met
Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser
Pro Gly Lys 210 215
220155220PRTArtificial SequenceVariant canine IgG-C Fc, Protein A +, C1q
, CD16 , D(38)G, K(93)R, K(97)I, A(98)G 155Pro Gly Cys Gly Leu Leu
Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro
Thr Val Thr Cys 20 25 30Val
Val Val Asp Leu Gly Pro Glu Asn Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Ser Lys Gln Val Gln Thr
Ala Asn Thr Gln Pro Arg Glu 50 55
60Glu Gln Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu
Ser Gly Lys Gln Phe Lys Cys Arg Val Asn Asn 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Glu Ile Ile
Ser Lys Thr Pro Gly 100 105
110Gln Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu
115 120 125Met Ser Lys Asn Thr Val Thr
Leu Thr Cys Leu Val Lys Asp Phe Phe 130 135
140Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Ile
Ser Leu Ser His Ser Pro Gly Lys 210 215
220156220PRTArtificial SequenceVariant canine IgG-C Fc, Protein A +, C1q
+, CD16 , L(5)P, P(39)R 156Pro Gly Cys Gly Pro Leu Gly Gly Pro Ser
Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys
20 25 30Val Val Val Asp Leu Asp
Arg Glu Asn Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro
Arg Glu 50 55 60Glu Gln Ser Asn Gly
Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65 70
75 80His Gln Asp Trp Leu Ser Gly Lys Gln Phe
Lys Cys Lys Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly
100 105 110Gln Ala His Gln Pro
Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val
Lys Asp Phe Phe 130 135 140Pro Pro Glu
Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Met Thr
Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser
Pro Gly Lys 210 215
220157220PRTArtificial SequenceVariant canine IgG-C Fc, Protein A +, C1q
. CD16 , M(5)P, P(39)R, K(93)R 157Pro Gly Cys Gly Pro Leu Gly Gly
Pro Ser Val Phe Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val
Thr Cys 20 25 30Val Val Val
Asp Leu Asp Arg Glu Asn Pro Glu Val Gln Ile Ser Trp 35
40 45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn
Thr Gln Pro Arg Glu 50 55 60Glu Gln
Ser Asn Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp Leu Ser Gly
Lys Gln Phe Lys Cys Arg Val Asn Asn 85 90
95Lys Ala Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys
Thr Pro Gly 100 105 110Gln Ala
His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg Asp Glu 115
120 125Met Ser Lys Asn Thr Val Thr Leu Thr Cys
Leu Val Lys Asp Phe Phe 130 135 140Pro
Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu Pro145
150 155 160Glu Ser Lys Tyr Arg Met
Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser 165
170 175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser
Arg Trp Gln Arg 180 185 190Gly
Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His Asn His 195
200 205Tyr Thr Gln Ile Ser Leu Ser His Ser
Pro Gly Lys 210 215
220158221PRTArtificial SequenceAglycosyl variant canine IgG A Fc,
N(68)XMISC_FEATURE(68)..(68)X = any amino acid except N 158Pro Val Pro
Glu Pro Leu Gly Gly Pro Ser Val Leu Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Arg Ile Thr
Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Glu Val
His Thr Ala Lys Thr Gln Ser Arg Glu 50 55
60Gln Gln Phe Xaa Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65
70 75 80His Gln Asp Trp
Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His 85
90 95Ile Asp Leu Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly 100 105
110Arg Ala His Lys Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu
115 120 125Leu Ser Ser Ser Asp Thr Val
Ser Ile Thr Cys Leu Ile Lys Asp Phe 130 135
140Tyr Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln
Glu145 150 155 160Pro Glu
Arg Lys His Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln 180 185
190Gln Gly Asp Pro Phe Thr Cys Ala Val Met His Glu Thr Leu
Gln Asn 195 200 205His Tyr Thr Asp
Leu Ser Leu Ser His Ser Pro Gly Lys 210 215
220159221PRTArtificial SequenceAglycosyl variant canine IgG A Fc,
G(69)P 159Pro Val Pro Glu Pro Leu Gly Gly Pro Ser Val Leu Ile Phe Pro
Pro1 5 10 15Lys Pro Lys
Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Val Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu Asp Pro
Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser Arg Glu 50
55 60Gln Gln Phe Asn Pro Thr Tyr Arg Val
Val Ser Val Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val
Asn His 85 90 95Ile Asp
Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Arg Ala His Lys Pro Ser Val Tyr Val
Leu Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Ser Ile Thr Cys Leu Ile Lys Asp Phe
130 135 140Tyr Pro Pro Asp Ile Asp Val
Glu Trp Gln Ser Asn Gly Gln Gln Glu145 150
155 160Pro Glu Arg Lys His Arg Met Thr Pro Pro Gln Leu
Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Pro Phe Thr
Cys Ala Val Met His Glu Thr Leu Gln Asn 195 200
205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
210 215 220160221PRTArtificial
SequenceAglycosyl variant canine IgG A Fc, T(70)XMISC_FEATURE(70)..(70)X
= any amino acid except T or S 160Pro Val Pro Glu Pro Leu Gly Gly Pro Ser
Val Leu Ile Phe Pro Pro1 5 10
15Lys Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Val Thr Cys
20 25 30Val Val Leu Asp Leu Gly
Arg Glu Asp Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Ser
Arg Glu 50 55 60Gln Gln Phe Asn Gly
Xaa Tyr Arg Val Val Ser Val Leu Pro Ile Glu65 70
75 80His Gln Asp Trp Leu Thr Gly Lys Glu Phe
Lys Cys Arg Val Asn His 85 90
95Ile Asp Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly
100 105 110Arg Ala His Lys Pro
Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu 115
120 125Leu Ser Ser Ser Asp Thr Val Ser Ile Thr Cys Leu
Ile Lys Asp Phe 130 135 140Tyr Pro Pro
Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu145
150 155 160Pro Glu Arg Lys His Arg Met
Thr Pro Pro Gln Leu Asp Glu Asp Gly 165
170 175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys
Ser Arg Trp Gln 180 185 190Gln
Gly Asp Pro Phe Thr Cys Ala Val Met His Glu Thr Leu Gln Asn 195
200 205His Tyr Thr Asp Leu Ser Leu Ser His
Ser Pro Gly Lys 210 215
220161220PRTArtificial SequenceAglycosyl variant canine IgG B Fc,
N(68)XMISC_FEATURE(68)..(68)X = any amino acid except N 161Pro Ala Pro
Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Thr Leu Leu Ile Ala
Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Gln Met
Gln Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Xaa Gly Thr Tyr Arg Val Val Ser Val Leu Pro Ile Gly65
70 75 80His Gln Asp Trp
Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn 85
90 95Lys Ala Leu Pro Ser Pro Ile Glu Arg Thr
Ile Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Arg Glu Glu
115 120 125Leu Ser Lys Asn Thr Val Ser
Leu Thr Cys Leu Ile Lys Asp Phe Phe 130 135
140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Gln Glu
Pro145 150 155 160Glu Ser
Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr Ser Lys Leu
Ser Val Asp Lys Ser Arg Trp Gln Arg 180 185
190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala Leu His
Asn His 195 200 205Tyr Thr Gln Glu
Ser Leu Ser His Ser Pro Gly Lys 210 215
220162220PRTArtificial SequenceAglycosyl variant canine IgG B Fc, G(69)P
162Pro Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Thr Leu
Leu Ile Ala Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln
Ile Ser Trp 35 40 45Phe Val Asp
Gly Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Glu Gln Phe Asn Pro Thr Tyr Arg Val Val Ser Val
Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn
85 90 95Lys Ala Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro
Ser Arg Glu Glu 115 120 125Leu Ser
Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp Phe Phe 130
135 140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210 215
220163220PRTArtificial SequenceAglycosyl variant canine IgG
B Fc, T(70)XMISC_FEATURE(70)..(70)X = any amino acid except T or S 163Pro
Ala Pro Glu Met Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1
5 10 15Lys Pro Lys Asp Thr Leu Leu
Ile Ala Arg Thr Pro Glu Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asp Pro Glu Val Gln Ile
Ser Trp 35 40 45Phe Val Asp Gly
Lys Gln Met Gln Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Glu Gln Phe Asn Gly Xaa Tyr Arg Val Val Ser Val Leu
Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Lys Gly Lys Gln Phe Thr Cys Lys Val Asn Asn
85 90 95Lys Ala Leu Pro Ser Pro
Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro
Ser Arg Glu Glu 115 120 125Leu Ser
Lys Asn Thr Val Ser Leu Thr Cys Leu Ile Lys Asp Phe Phe 130
135 140Pro Pro Asp Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Thr Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu
Ala Leu His Asn His 195 200 205Tyr
Thr Gln Glu Ser Leu Ser His Ser Pro Gly Lys 210 215
220164220PRTArtificial SequenceAglycosyl variant canine IgG
C Fc, N(68)XMISC_FEATURE(68)..(68)X = any amino acid except N 164Pro Gly
Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Val Thr
Ala Arg Thr Pro Thr Val Thr Cys 20 25
30Val Val Val Asp Leu Asp Pro Glu Asn Pro Glu Val Gln Ile Ser
Trp 35 40 45Phe Val Asp Ser Lys
Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50 55
60Glu Gln Ser Xaa Gly Thr Tyr Arg Val Val Ser Val Leu Pro
Ile Gly65 70 75 80His
Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val Asn Asn
85 90 95Lys Ala Leu Pro Ser Pro Ile
Glu Glu Ile Ile Ser Lys Thr Pro Gly 100 105
110Gln Ala His Gln Pro Asn Val Tyr Val Leu Pro Pro Ser Arg
Asp Glu 115 120 125Met Ser Lys Asn
Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe 130
135 140Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly
Gln Gln Glu Pro145 150 155
160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp Glu Asp Gly Ser
165 170 175Tyr Phe Leu Tyr Ser
Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg 180
185 190Gly Asp Thr Phe Ile Cys Ala Val Met His Glu Ala
Leu His Asn His 195 200 205Tyr Thr
Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210 215
220165220PRTArtificial SequenceAglycosyl variant canine IgG C
Fc, G(69)P 165Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro1 5 10 15Lys Pro Lys
Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu Asn Pro
Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50
55 60Glu Gln Ser Asn Pro Thr Tyr Arg Val
Val Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val
Asn Asn 85 90 95Lys Ala
Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly 100
105 110Gln Ala His Gln Pro Asn Val Tyr Val
Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe
130 135 140Pro Pro Glu Ile Asp Val Glu
Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp
Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210
215 220166220PRTArtificial SequenceAglycosyl
variant canine IgG C Fc, T(70)XMISC_FEATURE(70)..(70)X = any amino acid
except T or S 166Pro Gly Cys Gly Leu Leu Gly Gly Pro Ser Val Phe Ile Phe
Pro Pro1 5 10 15Lys Pro
Lys Asp Ile Leu Val Thr Ala Arg Thr Pro Thr Val Thr Cys 20
25 30Val Val Val Asp Leu Asp Pro Glu Asn
Pro Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Ser Lys Gln Val Gln Thr Ala Asn Thr Gln Pro Arg Glu 50
55 60Glu Gln Ser Asn Gly Xaa Tyr Arg Val
Val Ser Val Leu Pro Ile Gly65 70 75
80His Gln Asp Trp Leu Ser Gly Lys Gln Phe Lys Cys Lys Val
Asn Asn 85 90 95Lys Ala
Leu Pro Ser Pro Ile Glu Glu Ile Ile Ser Lys Thr Pro Gly 100
105 110Gln Ala His Gln Pro Asn Val Tyr Val
Leu Pro Pro Ser Arg Asp Glu 115 120
125Met Ser Lys Asn Thr Val Thr Leu Thr Cys Leu Val Lys Asp Phe Phe
130 135 140Pro Pro Glu Ile Asp Val Glu
Trp Gln Ser Asn Gly Gln Gln Glu Pro145 150
155 160Glu Ser Lys Tyr Arg Met Thr Pro Pro Gln Leu Asp
Glu Asp Gly Ser 165 170
175Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln Arg
180 185 190Gly Asp Thr Phe Ile Cys
Ala Val Met His Glu Ala Leu His Asn His 195 200
205Tyr Thr Gln Ile Ser Leu Ser His Ser Pro Gly Lys 210
215 220167221PRTArtificial SequenceAglycosyl
variant canine IgG D Fc, N(68)XMISC_FEATURE(68)..(68)X = any amino acid
except N 167Pro Val Pro Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro
Pro1 5 10 15Lys Pro Lys
Asp Ile Leu Arg Ile Thr Arg Thr Pro Glu Ile Thr Cys 20
25 30Val Val Leu Asp Leu Gly Arg Glu Asp Pro
Glu Val Gln Ile Ser Trp 35 40
45Phe Val Asp Gly Lys Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu 50
55 60Gln Gln Phe Xaa Ser Thr Tyr Arg Val
Val Ser Val Leu Pro Ile Glu65 70 75
80His Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val
Asn His 85 90 95Ile Gly
Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Arg Gly 100
105 110Gln Ala His Gln Pro Ser Val Tyr Val
Leu Pro Pro Ser Pro Lys Glu 115 120
125Leu Ser Ser Ser Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe
130 135 140Phe Pro Pro Glu Ile Asp Val
Glu Trp Gln Ser Asn Gly Gln Pro Glu145 150
155 160Pro Glu Ser Lys Tyr His Thr Thr Ala Pro Gln Leu
Asp Glu Asp Gly 165 170
175Ser Tyr Phe Leu Tyr Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln
180 185 190Gln Gly Asp Thr Phe Thr
Cys Ala Val Met His Glu Ala Leu Gln Asn 195 200
205His Tyr Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys
210 215 220168221PRTArtificial
SequenceAglycosyl variant canine IgG D Fc, S(69)P 168Pro Val Pro Glu Ser
Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Arg Ile Thr Arg Thr
Pro Glu Ile Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser Trp
35 40 45Phe Val Asp Gly Lys Glu Val His
Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Gln Gln Phe Asn Pro Thr Tyr Arg Val Val Ser Val Leu Pro Ile Glu65
70 75 80His Gln Asp Trp Leu
Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His 85
90 95Ile Gly Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro Lys Glu
115 120 125Leu Ser Ser Ser Asp Thr Val
Thr Leu Thr Cys Leu Ile Lys Asp Phe 130 135
140Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn Gly Gln Pro
Glu145 150 155 160Pro Glu
Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu Tyr Ser Lys
Leu Ser Val Asp Lys Ser Arg Trp Gln 180 185
190Gln Gly Asp Thr Phe Thr Cys Ala Val Met His Glu Ala Leu
Gln Asn 195 200 205His Tyr Thr Asp
Leu Ser Leu Ser His Ser Pro Gly Lys 210 215
220169221PRTArtificial SequenceAglycosyl variant canine IgG D Fc,
T(70)XMISC_FEATURE(70)..(70)X = any amino acid except T or S 169Pro Val
Pro Glu Ser Leu Gly Gly Pro Ser Val Phe Ile Phe Pro Pro1 5
10 15Lys Pro Lys Asp Ile Leu Arg Ile
Thr Arg Thr Pro Glu Ile Thr Cys 20 25
30Val Val Leu Asp Leu Gly Arg Glu Asp Pro Glu Val Gln Ile Ser
Trp 35 40 45Phe Val Asp Gly Lys
Glu Val His Thr Ala Lys Thr Gln Pro Arg Glu 50 55
60Gln Gln Phe Asn Ser Xaa Tyr Arg Val Val Ser Val Leu Pro
Ile Glu65 70 75 80His
Gln Asp Trp Leu Thr Gly Lys Glu Phe Lys Cys Arg Val Asn His
85 90 95Ile Gly Leu Pro Ser Pro Ile
Glu Arg Thr Ile Ser Lys Ala Arg Gly 100 105
110Gln Ala His Gln Pro Ser Val Tyr Val Leu Pro Pro Ser Pro
Lys Glu 115 120 125Leu Ser Ser Ser
Asp Thr Val Thr Leu Thr Cys Leu Ile Lys Asp Phe 130
135 140Phe Pro Pro Glu Ile Asp Val Glu Trp Gln Ser Asn
Gly Gln Pro Glu145 150 155
160Pro Glu Ser Lys Tyr His Thr Thr Ala Pro Gln Leu Asp Glu Asp Gly
165 170 175Ser Tyr Phe Leu Tyr
Ser Lys Leu Ser Val Asp Lys Ser Arg Trp Gln 180
185 190Gln Gly Asp Thr Phe Thr Cys Ala Val Met His Glu
Ala Leu Gln Asn 195 200 205His Tyr
Thr Asp Leu Ser Leu Ser His Ser Pro Gly Lys 210 215
220170237PRTArtificial SequenceAglycosyl variant feline
IgG1a Fc, N(85)XMISC_FEATURE(85)..(85)X = any amino acid except N 170Arg
Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1
5 10 15Cys Pro Pro Pro Glu Met Leu
Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu
Val Thr 35 40 45Cys Leu Val Val
Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr
Ser Pro Arg65 70 75
80Glu Glu Gln Phe Xaa Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Ser Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Val Tyr Ser Lys Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235171237PRTArtificial
SequenceAglycosyl variant feline IgG1a Fc, S(86)P 171Arg Lys Thr Asp His
Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1 5
10 15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser
Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
35 40 45Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65
70 75 80Glu Glu Gln Phe Asn
Pro Thr Tyr Arg Val Val Ser Val Leu Pro Ile 85
90 95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe
Lys Cys Lys Val Asn 100 105
110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Lys
115 120 125Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro Pro Ala Gln Glu 130 135
140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Ser
Phe145 150 155 160His Pro
Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn Tyr Arg Thr
Thr Pro Pro Gln Leu Asp Ser Asp Gly 180 185
190Thr Tyr Phe Val Tyr Ser Lys Leu Ser Val Asp Arg Ser His
Trp Gln 195 200 205Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly
Lys225 230 235172237PRTArtificial
SequenceAglycosyl variant feline IgG1a Fc, T(87)XMISC_FEATURE(87)..(87)X
= any amino acid except T or S 172Arg Lys Thr Asp His Pro Pro Gly Pro Lys
Pro Cys Asp Cys Pro Lys1 5 10
15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro
20 25 30Pro Lys Pro Lys Asp Thr
Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln
Ile Thr 50 55 60Trp Phe Val Asp Asn
Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65 70
75 80Glu Glu Gln Phe Asn Ser Xaa Tyr Arg Val
Val Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn
100 105 110Ser Lys Ser Leu Pro
Ser Pro Ile Glu Arg Thr Ile Ser Lys Ala Lys 115
120 125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro
Pro Ala Gln Glu 130 135 140Glu Leu Ser
Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Ser Phe145
150 155 160His Pro Pro Asp Ile Ala Val
Glu Trp Glu Ile Thr Gly Gln Pro Glu 165
170 175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly 180 185 190Thr
Tyr Phe Val Tyr Ser Lys Leu Ser Val Asp Arg Ser His Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His Ser 210 215
220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225
230 235173237PRTArtificial SequenceAglycosyl variant
feline IgG1b Fc, N(85)XMISC_FEATURE(85)..(85)X = any amino acid except N
173Arg Lys Thr Asp His Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1
5 10 15Cys Pro Pro Pro Glu Met
Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro
Glu Val Thr 35 40 45Cys Leu Val
Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln Ile Thr 50
55 60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys
Thr Ser Pro Arg65 70 75
80Glu Glu Gln Phe Xaa Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile
Ser Lys Asp Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Ala Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Glu Gly Phe145 150 155
160Tyr Pro Ser Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Arg Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser Arg Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235174237PRTArtificial
SequenceAglycosyl variant feline IgG1b Fc, S(86)P 174Arg Lys Thr Asp His
Pro Pro Gly Pro Lys Pro Cys Asp Cys Pro Lys1 5
10 15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser
Ile Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
35 40 45Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asp Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65
70 75 80Glu Glu Gln Phe Asn
Pro Thr Tyr Arg Val Val Ser Val Leu Pro Ile 85
90 95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe
Lys Cys Lys Val Asn 100 105
110Ser Lys Ser Leu Pro Ser Pro Ile Glu Arg Thr Ile Ser Lys Asp Lys
115 120 125Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro Pro Ala Gln Glu 130 135
140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Glu Gly
Phe145 150 155 160Tyr Pro
Ser Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn Tyr Arg Thr
Thr Pro Pro Gln Leu Asp Ser Asp Gly 180 185
190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser Arg
Trp Gln 195 200 205Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly
Lys225 230 235175237PRTArtificial
SequenceAglycosyl variant feline IgG1b Fc, T(87)XMISC_FEATURE(87)..(87)X
= any amino acid except T or S 175Arg Lys Thr Asp His Pro Pro Gly Pro Lys
Pro Cys Asp Cys Pro Lys1 5 10
15Cys Pro Pro Pro Glu Met Leu Gly Gly Pro Ser Ile Phe Ile Phe Pro
20 25 30Pro Lys Pro Lys Asp Thr
Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asp Val Gln
Ile Thr 50 55 60Trp Phe Val Asp Asn
Thr Gln Val Tyr Thr Ala Lys Thr Ser Pro Arg65 70
75 80Glu Glu Gln Phe Asn Ser Xaa Tyr Arg Val
Val Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn
100 105 110Ser Lys Ser Leu Pro
Ser Pro Ile Glu Arg Thr Ile Ser Lys Asp Lys 115
120 125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro
Pro Ala Gln Glu 130 135 140Glu Leu Ser
Glu Asn Lys Val Ser Val Thr Cys Leu Ile Glu Gly Phe145
150 155 160Tyr Pro Ser Asp Ile Ala Val
Glu Trp Glu Ile Thr Gly Gln Pro Glu 165
170 175Pro Glu Asn Asn Tyr Arg Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly 180 185 190Thr
Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser Arg Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His Ser 210 215
220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225
230 235176237PRTArtificial SequenceAglycosyl variant
feline IgG2 Fc, N(85)XMISC_FEATURE(85)..(85)X= any amino acid except N
176Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Lys1
5 10 15Cys Pro Val Pro Glu Ile
Pro Gly Ala Pro Ser Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro
Glu Val Thr 35 40 45Cys Leu Val
Val Asp Leu Gly Pro Asp Asp Ser Asn Val Gln Ile Thr 50
55 60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys
Thr Arg Pro Arg65 70 75
80Glu Glu Gln Phe Xaa Ser Thr Tyr Arg Val Val Ser Val Leu Pro Ile
85 90 95Leu His Gln Asp Trp Leu
Lys Gly Lys Glu Phe Lys Cys Lys Val Asn 100
105 110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile
Ser Lys Ala Lys 115 120 125Gly Gln
Pro His Glu Pro Gln Val Tyr Val Leu Pro Pro Thr Gln Glu 130
135 140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys
Leu Ile Lys Gly Phe145 150 155
160His Pro Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn
Tyr Gln Thr Thr Pro Pro Gln Leu Asp Ser Asp Gly 180
185 190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp
Arg Ser His Trp Gln 195 200 205Arg
Gly Asn Thr Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser
Pro Gly Lys225 230 235177237PRTArtificial
SequenceAglycosyl variant feline IgG2 Fc, S(86)P 177Pro Lys Thr Ala Ser
Thr Ile Glu Ser Lys Thr Gly Glu Gly Pro Lys1 5
10 15Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser
Val Phe Ile Phe Pro 20 25
30Pro Lys Pro Lys Asp Thr Leu Ser Ile Ser Arg Thr Pro Glu Val Thr
35 40 45Cys Leu Val Val Asp Leu Gly Pro
Asp Asp Ser Asn Val Gln Ile Thr 50 55
60Trp Phe Val Asp Asn Thr Glu Met His Thr Ala Lys Thr Arg Pro Arg65
70 75 80Glu Glu Gln Phe Asn
Pro Thr Tyr Arg Val Val Ser Val Leu Pro Ile 85
90 95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe
Lys Cys Lys Val Asn 100 105
110Ser Lys Ser Leu Pro Ser Ala Met Glu Arg Thr Ile Ser Lys Ala Lys
115 120 125Gly Gln Pro His Glu Pro Gln
Val Tyr Val Leu Pro Pro Thr Gln Glu 130 135
140Glu Leu Ser Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Gly
Phe145 150 155 160His Pro
Pro Asp Ile Ala Val Glu Trp Glu Ile Thr Gly Gln Pro Glu
165 170 175Pro Glu Asn Asn Tyr Gln Thr
Thr Pro Pro Gln Leu Asp Ser Asp Gly 180 185
190Thr Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His
Trp Gln 195 200 205Arg Gly Asn Thr
Tyr Thr Cys Ser Val Ser His Glu Ala Leu His Ser 210
215 220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly
Lys225 230 235178237PRTArtificial
SequenceAglycosyl variant feline IgG2 Fc, T(87)XMISC_FEATURE(87)..(87)X =
any amino acid except T or S 178Pro Lys Thr Ala Ser Thr Ile Glu Ser Lys
Thr Gly Glu Gly Pro Lys1 5 10
15Cys Pro Val Pro Glu Ile Pro Gly Ala Pro Ser Val Phe Ile Phe Pro
20 25 30Pro Lys Pro Lys Asp Thr
Leu Ser Ile Ser Arg Thr Pro Glu Val Thr 35 40
45Cys Leu Val Val Asp Leu Gly Pro Asp Asp Ser Asn Val Gln
Ile Thr 50 55 60Trp Phe Val Asp Asn
Thr Glu Met His Thr Ala Lys Thr Arg Pro Arg65 70
75 80Glu Glu Gln Phe Asn Ser Xaa Tyr Arg Val
Val Ser Val Leu Pro Ile 85 90
95Leu His Gln Asp Trp Leu Lys Gly Lys Glu Phe Lys Cys Lys Val Asn
100 105 110Ser Lys Ser Leu Pro
Ser Ala Met Glu Arg Thr Ile Ser Lys Ala Lys 115
120 125Gly Gln Pro His Glu Pro Gln Val Tyr Val Leu Pro
Pro Thr Gln Glu 130 135 140Glu Leu Ser
Glu Asn Lys Val Ser Val Thr Cys Leu Ile Lys Gly Phe145
150 155 160His Pro Pro Asp Ile Ala Val
Glu Trp Glu Ile Thr Gly Gln Pro Glu 165
170 175Pro Glu Asn Asn Tyr Gln Thr Thr Pro Pro Gln Leu
Asp Ser Asp Gly 180 185 190Thr
Tyr Phe Leu Tyr Ser Arg Leu Ser Val Asp Arg Ser His Trp Gln 195
200 205Arg Gly Asn Thr Tyr Thr Cys Ser Val
Ser His Glu Ala Leu His Ser 210 215
220His His Thr Gln Lys Ser Leu Thr Gln Ser Pro Gly Lys225
230 235179214PRTArtificial SequenceAglycosyl variant
equine IgG1 Fc, N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N
179Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Asn Pro Lys Asp Thr Leu1
5 10 15Met Ile Thr Arg Thr Pro
Glu Val Thr Cys Val Val Val Asp Val Ser 20 25
30Gln Glu Asn Pro Asp Val Lys Phe Asn Trp Tyr Met Asp
Gly Val Glu 35 40 45Val Arg Thr
Ala Thr Thr Arg Pro Lys Glu Glu Gln Phe Xaa Ser Thr 50
55 60Tyr Arg Val Val Ser Val Leu Arg Ile Gln His Gln
Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Gln Ala Leu Pro Gln Pro
85 90 95Ile Glu Arg Thr Ile Thr
Lys Thr Lys Gly Arg Ser Gln Glu Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Pro Asp Glu Ser Lys
Lys Ser Lys Val 115 120 125Ser Val
Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Glu Ile Asn Ile 130
135 140Glu Trp Gln Ser Asn Gly Gln Pro Glu Leu Glu
Thr Lys Tyr Ser Thr145 150 155
160Thr Gln Ala Gln Gln Asp Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Ser Val Asp
Arg Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys 180
185 190Gly Val Met His Glu Ala Leu His Asn His Tyr
Thr Gln Lys Asn Val 195 200 205Ser
Lys Asn Pro Gly Lys 210180214PRTArtificial SequenceAglycosyl variant
equine IgG1 Fc, S(63)P 180Gly Gly Pro Ser Val Phe Leu Phe Pro Pro Asn Pro
Lys Asp Thr Leu1 5 10
15Met Ile Thr Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
20 25 30Gln Glu Asn Pro Asp Val Lys
Phe Asn Trp Tyr Met Asp Gly Val Glu 35 40
45Val Arg Thr Ala Thr Thr Arg Pro Lys Glu Glu Gln Phe Asn Pro
Thr 50 55 60Tyr Arg Val Val Ser Val
Leu Arg Ile Gln His Gln Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Gln
Ala Leu Pro Gln Pro 85 90
95Ile Glu Arg Thr Ile Thr Lys Thr Lys Gly Arg Ser Gln Glu Pro Gln
100 105 110Val Tyr Val Leu Ala Pro
His Pro Asp Glu Ser Lys Lys Ser Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Glu Ile
Asn Ile 130 135 140Glu Trp Gln Ser Asn
Gly Gln Pro Glu Leu Glu Thr Lys Tyr Ser Thr145 150
155 160Thr Gln Ala Gln Gln Asp Ser Asp Gly Ser
Tyr Phe Leu Tyr Ser Lys 165 170
175Leu Ser Val Asp Arg Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Gly Val Met His Glu
Ala Leu His Asn His Tyr Thr Gln Lys Asn Val 195
200 205Ser Lys Asn Pro Gly Lys 210181214PRTArtificial
SequenceAglycosyl variant equine IgG1 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 181Gly Gly Pro Ser Val Phe Leu Phe Pro Pro
Asn Pro Lys Asp Thr Leu1 5 10
15Met Ile Thr Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
20 25 30Gln Glu Asn Pro Asp Val
Lys Phe Asn Trp Tyr Met Asp Gly Val Glu 35 40
45Val Arg Thr Ala Thr Thr Arg Pro Lys Glu Glu Gln Phe Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Val Leu Arg Ile Gln His Gln Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn
Gln Ala Leu Pro Gln Pro 85 90
95Ile Glu Arg Thr Ile Thr Lys Thr Lys Gly Arg Ser Gln Glu Pro Gln
100 105 110Val Tyr Val Leu Ala
Pro His Pro Asp Glu Ser Lys Lys Ser Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Glu Ile Asn Ile 130 135 140Glu Trp Gln
Ser Asn Gly Gln Pro Glu Leu Glu Thr Lys Tyr Ser Thr145
150 155 160Thr Gln Ala Gln Gln Asp Ser
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Ser Val Asp Arg Asn Arg Trp Gln Gln Gly Thr
Thr Phe Thr Cys 180 185 190Gly
Val Met His Glu Ala Leu His Asn His Tyr Thr Gln Lys Asn Val 195
200 205Ser Lys Asn Pro Gly Lys
210182214PRTArtificial SequenceAglycosyl variant equine IgG2 Fc,
N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N 182Gly Gly Pro
Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala Leu1 5
10 15Met Ile Ser Arg Thr Pro Val Val Thr
Cys Val Val Val Asn Leu Ser 20 25
30Asp Gln Tyr Pro Asp Val Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu
35 40 45Val His Ser Ala Ile Thr Lys
Gln Arg Glu Ala Gln Phe Xaa Ser Thr 50 55
60Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser65
70 75 80Gly Lys Glu Phe
Lys Cys Ser Val Thr Asn Val Gly Val Pro Gln Pro 85
90 95Ile Ser Arg Ala Ile Ser Arg Gly Lys Gly
Pro Ser Arg Val Pro Gln 100 105
110Val Tyr Val Leu Pro Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val
115 120 125Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Ser Val 130 135
140Glu Trp Gln Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser
Thr145 150 155 160Thr Pro
Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Ser Leu Glu Thr Ser Arg
Trp Gln Gln Val Glu Ser Phe Thr Cys 180 185
190Ala Val Met His Glu Ala Leu His Asn His Phe Thr Lys Thr
Asp Ile 195 200 205Ser Glu Ser Leu
Gly Lys 210183214PRTArtificial SequenceAglycosyl variant equine IgG2
Fc, S(63)P 183Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Ala
Leu1 5 10 15Met Ile Ser
Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser 20
25 30Asp Gln Tyr Pro Asp Val Gln Phe Ser Trp
Tyr Val Asp Asn Thr Glu 35 40
45Val His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn Pro Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Thr Asn Val Gly Val Pro
Gln Pro 85 90 95Ile Ser
Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln 100
105 110Val Tyr Val Leu Pro Pro His Pro Asp
Glu Leu Ala Lys Ser Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Ser Val
130 135 140Glu Trp Gln Ser Asn Arg Trp
Pro Glu Leu Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu Ser Phe Thr Cys
180 185 190Ala Val Met His Glu Ala
Leu His Asn His Phe Thr Lys Thr Asp Ile 195 200
205Ser Glu Ser Leu Gly Lys 210184214PRTArtificial
SequenceAglycosyl variant equine IgG2 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 184Gly Gly Pro Ser Val Phe Ile Phe Pro Pro
Asn Pro Lys Asp Ala Leu1 5 10
15Met Ile Ser Arg Thr Pro Val Val Thr Cys Val Val Val Asn Leu Ser
20 25 30Asp Gln Tyr Pro Asp Val
Gln Phe Ser Trp Tyr Val Asp Asn Thr Glu 35 40
45Val His Ser Ala Ile Thr Lys Gln Arg Glu Ala Gln Phe Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Ser Val Thr Asn
Val Gly Val Pro Gln Pro 85 90
95Ile Ser Arg Ala Ile Ser Arg Gly Lys Gly Pro Ser Arg Val Pro Gln
100 105 110Val Tyr Val Leu Pro
Pro His Pro Asp Glu Leu Ala Lys Ser Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Asp Ile Ser Val 130 135 140Glu Trp Gln
Ser Asn Arg Trp Pro Glu Leu Glu Gly Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asp Gly
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Ser Leu Glu Thr Ser Arg Trp Gln Gln Val Glu
Ser Phe Thr Cys 180 185 190Ala
Val Met His Glu Ala Leu His Asn His Phe Thr Lys Thr Asp Ile 195
200 205Ser Glu Ser Leu Gly Lys
210185214PRTArtificial SequenceAglycosyl variant equine IgG3 Fc,
N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N 185Gly Gly Pro
Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1 5
10 15Met Ile Thr Arg Met Pro Glu Val Thr
Cys Leu Val Val Asp Val Ser 20 25
30His Asp Ser Ser Asp Val Leu Phe Thr Trp Tyr Val Asp Gly Thr Glu
35 40 45Val Lys Thr Ala Lys Thr Met
Pro Asn Glu Glu Gln Asn Xaa Ser Thr 50 55
60Tyr Arg Val Val Ser Val Leu Arg Ile Gln His Gln Asp Trp Leu Asn65
70 75 80Gly Lys Lys Phe
Lys Cys Lys Val Asn Asn Gln Ala Leu Pro Ala Pro 85
90 95Val Glu Arg Thr Ile Ser Lys Ala Thr Gly
Gln Thr Arg Val Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Pro Asp Glu Leu Ser Lys Asn Lys Val
115 120 125Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Thr Val 130 135
140Glu Trp Gln Ser Asn Glu His Pro Glu Pro Glu Gly Lys Tyr Arg
Thr145 150 155 160Thr Glu
Ala Gln Lys Asp Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Thr Val Glu Lys Asp Arg
Trp Gln Gln Gly Thr Thr Phe Thr Cys 180 185
190Val Val Met His Glu Ala Leu His Asn His Val Met Gln Lys
Asn Ile 195 200 205Ser Lys Asn Pro
Gly Lys 210186214PRTArtificial SequenceAglycosyl variant equine IgG3
Fc, S(63)P 186Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val
Leu1 5 10 15Met Ile Thr
Arg Met Pro Glu Val Thr Cys Leu Val Val Asp Val Ser 20
25 30His Asp Ser Ser Asp Val Leu Phe Thr Trp
Tyr Val Asp Gly Thr Glu 35 40
45Val Lys Thr Ala Lys Thr Met Pro Asn Glu Glu Gln Asn Asn Pro Thr 50
55 60Tyr Arg Val Val Ser Val Leu Arg Ile
Gln His Gln Asp Trp Leu Asn65 70 75
80Gly Lys Lys Phe Lys Cys Lys Val Asn Asn Gln Ala Leu Pro
Ala Pro 85 90 95Val Glu
Arg Thr Ile Ser Lys Ala Thr Gly Gln Thr Arg Val Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Pro Asp
Glu Leu Ser Lys Asn Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Thr Val
130 135 140Glu Trp Gln Ser Asn Glu His
Pro Glu Pro Glu Gly Lys Tyr Arg Thr145 150
155 160Thr Glu Ala Gln Lys Asp Ser Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Lys Asp Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Val Val Met His Glu Ala
Leu His Asn His Val Met Gln Lys Asn Ile 195 200
205Ser Lys Asn Pro Gly Lys 210187214PRTArtificial
SequenceAglycosyl variant equine IgG3 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 187Gly Gly Pro Ser Val Phe Ile Phe Pro Pro
Lys Pro Lys Asp Val Leu1 5 10
15Met Ile Thr Arg Met Pro Glu Val Thr Cys Leu Val Val Asp Val Ser
20 25 30His Asp Ser Ser Asp Val
Leu Phe Thr Trp Tyr Val Asp Gly Thr Glu 35 40
45Val Lys Thr Ala Lys Thr Met Pro Asn Glu Glu Gln Asn Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Val Leu Arg Ile Gln His Gln Asp Trp Leu Asn65 70
75 80Gly Lys Lys Phe Lys Cys Lys Val Asn Asn
Gln Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Ser Lys Ala Thr Gly Gln Thr Arg Val Pro Gln
100 105 110Val Tyr Val Leu Ala
Pro His Pro Asp Glu Leu Ser Lys Asn Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Asp Ile Thr Val 130 135 140Glu Trp Gln
Ser Asn Glu His Pro Glu Pro Glu Gly Lys Tyr Arg Thr145
150 155 160Thr Glu Ala Gln Lys Asp Ser
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Lys Asp Arg Trp Gln Gln Gly Thr
Thr Phe Thr Cys 180 185 190Val
Val Met His Glu Ala Leu His Asn His Val Met Gln Lys Asn Ile 195
200 205Ser Lys Asn Pro Gly Lys
210188214PRTArtificial SequenceAglycosyl variant equine IgG4 Fc,
N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N 188Val Gly Pro
Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1 5
10 15Met Ile Ser Arg Thr Pro Thr Val Thr
Cys Val Val Val Asp Val Gly 20 25
30His Asp Phe Pro Asp Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu
35 40 45Thr His Thr Ala Thr Thr Glu
Pro Lys Gln Glu Gln Phe Xaa Ser Thr 50 55
60Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Lys Asp Trp Leu Ser65
70 75 80Gly Lys Glu Phe
Lys Cys Lys Val Asn Asn Lys Ala Leu Pro Ala Pro 85
90 95Val Glu Arg Thr Ile Ser Ala Pro Thr Gly
Gln Pro Arg Glu Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Arg Asp Glu Leu Ser Lys Asn Lys Val
115 120 125Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Asp Ile 130 135
140Glu Trp Lys Ser Asn Gly Gln Pro Glu Pro Glu Thr Lys Tyr Ser
Thr145 150 155 160Thr Pro
Ala Gln Leu Asp Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Thr Val Glu Thr Asn Arg
Trp Gln Gln Gly Thr Thr Phe Thr Cys 180 185
190Ala Val Met His Glu Ala Leu His Asn His Tyr Thr Glu Lys
Ser Val 195 200 205Ser Lys Ser Pro
Gly Lys 210189214PRTArtificial SequenceAglycosyl variant equine IgG4
Fc, S(63)P 189Val Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val
Leu1 5 10 15Met Ile Ser
Arg Thr Pro Thr Val Thr Cys Val Val Val Asp Val Gly 20
25 30His Asp Phe Pro Asp Val Gln Phe Asn Trp
Tyr Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Gln Glu Gln Phe Asn Pro Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Lys Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Lys Ala Leu Pro
Ala Pro 85 90 95Val Glu
Arg Thr Ile Ser Ala Pro Thr Gly Gln Pro Arg Glu Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Arg Asp
Glu Leu Ser Lys Asn Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Asp Ile
130 135 140Glu Trp Lys Ser Asn Gly Gln
Pro Glu Pro Glu Thr Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Ser Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Thr Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr Glu Lys Ser Val 195 200
205Ser Lys Ser Pro Gly Lys 210190214PRTArtificial
SequenceAglycosyl variant equine IgG4 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 190Val Gly Pro Ser Val Phe Ile Phe Pro Pro
Lys Pro Lys Asp Val Leu1 5 10
15Met Ile Ser Arg Thr Pro Thr Val Thr Cys Val Val Val Asp Val Gly
20 25 30His Asp Phe Pro Asp Val
Gln Phe Asn Trp Tyr Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Gln Glu Gln Phe Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Val Leu Pro Ile Gln His Lys Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn
Lys Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Ser Ala Pro Thr Gly Gln Pro Arg Glu Pro Gln
100 105 110Val Tyr Val Leu Ala
Pro His Arg Asp Glu Leu Ser Lys Asn Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Asp Ile Asp Ile 130 135 140Glu Trp Lys
Ser Asn Gly Gln Pro Glu Pro Glu Thr Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asp Ser
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Thr Asn Arg Trp Gln Gln Gly Thr
Thr Phe Thr Cys 180 185 190Ala
Val Met His Glu Ala Leu His Asn His Tyr Thr Glu Lys Ser Val 195
200 205Ser Lys Ser Pro Gly Lys
210191214PRTArtificial SequenceAglycosyl variant equine IgG5 Fc,
N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N 191Gly Gly Pro
Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1 5
10 15Met Ile Ser Arg Lys Pro Glu Val Thr
Cys Val Val Val Asp Leu Gly 20 25
30His Asp Asp Pro Asp Val Gln Phe Thr Trp Phe Val Asp Gly Val Glu
35 40 45Thr His Thr Ala Thr Thr Glu
Pro Lys Glu Glu Gln Phe Xaa Pro Thr 50 55
60Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser65
70 75 80Gly Lys Glu Phe
Lys Cys Ser Val Thr Ser Lys Ala Leu Pro Ala Pro 85
90 95Val Glu Arg Thr Ile Ser Lys Ala Lys Gly
Gln Leu Arg Val Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Pro Asp Glu Leu Ala Lys Asn Thr Val
115 120 125Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Glu Ile Asp Val 130 135
140Glu Trp Gln Ser Asn Glu His Pro Glu Pro Glu Gly Lys Tyr Ser
Thr145 150 155 160Thr Pro
Ala Gln Leu Asn Ser Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Ser Val Glu Thr Ser Arg
Trp Lys Gln Gly Glu Ser Phe Thr Cys 180 185
190Gly Val Met His Glu Ala Val Glu Asn His Tyr Thr Gln Lys
Asn Val 195 200 205Ser His Ser Pro
Gly Lys 210192214PRTArtificial SequenceAglycosyl variant equine IgG5
Fc, S(63)P 192Gly Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val
Leu1 5 10 15Met Ile Ser
Arg Lys Pro Glu Val Thr Cys Val Val Val Asp Leu Gly 20
25 30His Asp Asp Pro Asp Val Gln Phe Thr Trp
Phe Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Glu Glu Gln Phe Asn Pro Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Ser Val Thr Ser Lys Ala Leu Pro
Ala Pro 85 90 95Val Glu
Arg Thr Ile Ser Lys Ala Lys Gly Gln Leu Arg Val Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Pro Asp
Glu Leu Ala Lys Asn Thr Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Glu Ile Asp Val
130 135 140Glu Trp Gln Ser Asn Glu His
Pro Glu Pro Glu Gly Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asn Ser Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Ser Val Glu Thr Ser Arg Trp Lys Gln Gly Glu Ser Phe Thr Cys
180 185 190Gly Val Met His Glu Ala
Val Glu Asn His Tyr Thr Gln Lys Asn Val 195 200
205Ser His Ser Pro Gly Lys 210193214PRTArtificial
SequenceAglycosyl variant equine IgG5 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 193Gly Gly Pro Ser Val Phe Ile Phe Pro Pro
Lys Pro Lys Asp Val Leu1 5 10
15Met Ile Ser Arg Lys Pro Glu Val Thr Cys Val Val Val Asp Leu Gly
20 25 30His Asp Asp Pro Asp Val
Gln Phe Thr Trp Phe Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Glu Glu Gln Phe Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Val Leu Pro Ile Gln His Gln Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Ser Val Thr Ser
Lys Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Ser Lys Ala Lys Gly Gln Leu Arg Val Pro Gln
100 105 110Val Tyr Val Leu Ala
Pro His Pro Asp Glu Leu Ala Lys Asn Thr Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Glu Ile Asp Val 130 135 140Glu Trp Gln
Ser Asn Glu His Pro Glu Pro Glu Gly Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asn Ser
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Ser Val Glu Thr Ser Arg Trp Lys Gln Gly Glu
Ser Phe Thr Cys 180 185 190Gly
Val Met His Glu Ala Val Glu Asn His Tyr Thr Gln Lys Asn Val 195
200 205Ser His Ser Pro Gly Lys
210194214PRTArtificial SequenceAglycosyl variant equine IgG6 Fc,
N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N 194Gly Arg Pro
Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Thr Leu1 5
10 15Met Ile Ser Arg Thr Pro Glu Val Thr
Cys Val Val Val Asp Val Ser 20 25
30Gln Glu Asn Pro Asp Val Lys Phe Asn Trp Tyr Val Asp Gly Val Glu
35 40 45Ala His Thr Ala Thr Thr Lys
Ala Lys Glu Lys Gln Asp Xaa Ser Thr 50 55
60Tyr Arg Val Val Ser Val Leu Pro Ile Gln His Gln Asp Trp Arg Arg65
70 75 80Gly Lys Glu Phe
Lys Cys Lys Val Asn Asn Arg Ala Leu Pro Ala Pro 85
90 95Val Glu Arg Thr Ile Thr Lys Ala Lys Gly
Glu Leu Gln Asp Pro Gln 100 105
110Val Tyr Ile Leu Ala Pro His Pro Asp Glu Val Thr Lys Asn Thr Val
115 120 125Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Asn Val 130 135
140Glu Trp Gln Ser Asn Glu Glu Pro Glu Pro Glu Val Lys Tyr Ser
Thr145 150 155 160Thr Pro
Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Thr Val Glu Thr Asp Arg
Trp Glu Gln Gly Glu Ser Phe Thr Cys 180 185
190Val Val Met His Glu Ala Ile Arg His Thr Tyr Arg Gln Lys
Ser Ile 195 200 205Thr Asn Phe Pro
Gly Lys 210195214PRTArtificial SequenceAglycosyl variant equine IgG6
Fc, S(63)P 195Gly Arg Pro Ser Val Phe Ile Phe Pro Pro Asn Pro Lys Asp Thr
Leu1 5 10 15Met Ile Ser
Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser 20
25 30Gln Glu Asn Pro Asp Val Lys Phe Asn Trp
Tyr Val Asp Gly Val Glu 35 40
45Ala His Thr Ala Thr Thr Lys Ala Lys Glu Lys Gln Asp Asn Pro Thr 50
55 60Tyr Arg Val Val Ser Val Leu Pro Ile
Gln His Gln Asp Trp Arg Arg65 70 75
80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Arg Ala Leu Pro
Ala Pro 85 90 95Val Glu
Arg Thr Ile Thr Lys Ala Lys Gly Glu Leu Gln Asp Pro Gln 100
105 110Val Tyr Ile Leu Ala Pro His Pro Asp
Glu Val Thr Lys Asn Thr Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Asn Val
130 135 140Glu Trp Gln Ser Asn Glu Glu
Pro Glu Pro Glu Val Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Thr Asp Arg Trp Glu Gln Gly Glu Ser Phe Thr Cys
180 185 190Val Val Met His Glu Ala
Ile Arg His Thr Tyr Arg Gln Lys Ser Ile 195 200
205Thr Asn Phe Pro Gly Lys 210196214PRTArtificial
SequenceAglycosyl variant equine IgG6 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 196Gly Arg Pro Ser Val Phe Ile Phe Pro Pro
Asn Pro Lys Asp Thr Leu1 5 10
15Met Ile Ser Arg Thr Pro Glu Val Thr Cys Val Val Val Asp Val Ser
20 25 30Gln Glu Asn Pro Asp Val
Lys Phe Asn Trp Tyr Val Asp Gly Val Glu 35 40
45Ala His Thr Ala Thr Thr Lys Ala Lys Glu Lys Gln Asp Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Val Leu Pro Ile Gln His Gln Asp Trp Arg Arg65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn
Arg Ala Leu Pro Ala Pro 85 90
95Val Glu Arg Thr Ile Thr Lys Ala Lys Gly Glu Leu Gln Asp Pro Gln
100 105 110Val Tyr Ile Leu Ala
Pro His Pro Asp Glu Val Thr Lys Asn Thr Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Asp Ile Asn Val 130 135 140Glu Trp Gln
Ser Asn Glu Glu Pro Glu Pro Glu Val Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asp Gly
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Thr Asp Arg Trp Glu Gln Gly Glu
Ser Phe Thr Cys 180 185 190Val
Val Met His Glu Ala Ile Arg His Thr Tyr Arg Gln Lys Ser Ile 195
200 205Thr Asn Phe Pro Gly Lys
210197214PRTArtificial SequenceAglycosyl variant equine IgG7 Fc,
N(62)XMISC_FEATURE(62)..(62)X = any amino acid except N 197Val Gly Pro
Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val Leu1 5
10 15Met Ile Ser Arg Thr Pro Thr Val Thr
Cys Val Val Val Asp Val Gly 20 25
30His Asp Phe Pro Asp Val Gln Phe Asn Trp Tyr Val Asp Gly Val Glu
35 40 45Thr His Thr Ala Thr Thr Glu
Pro Lys Gln Glu Gln Asn Xaa Ser Thr 50 55
60Tyr Arg Val Val Ser Ile Leu Ala Ile Gln His Lys Asp Trp Leu Ser65
70 75 80Gly Lys Glu Phe
Lys Cys Lys Val Asn Asn Gln Ala Leu Pro Ala Pro 85
90 95Val Gln Lys Thr Ile Ser Lys Pro Thr Gly
Gln Pro Arg Glu Pro Gln 100 105
110Val Tyr Val Leu Ala Pro His Pro Asp Glu Leu Ser Lys Asn Lys Val
115 120 125Ser Val Thr Cys Leu Val Lys
Asp Phe Tyr Pro Pro Asp Ile Asp Ile 130 135
140Glu Trp Lys Ser Asn Gly Gln Pro Glu Pro Glu Thr Lys Tyr Ser
Thr145 150 155 160Thr Pro
Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe Leu Tyr Ser Lys
165 170 175Leu Thr Val Glu Thr Asn Arg
Trp Gln Gln Gly Thr Thr Phe Thr Cys 180 185
190Ala Val Met His Glu Ala Leu His Asn His Tyr Thr Glu Lys
Ser Val 195 200 205Ser Lys Ser Pro
Gly Lys 210198214PRTArtificial SequenceAglycosyl variant equine IgG7
Fc, S(63)P 198Val Gly Pro Ser Val Phe Ile Phe Pro Pro Lys Pro Lys Asp Val
Leu1 5 10 15Met Ile Ser
Arg Thr Pro Thr Val Thr Cys Val Val Val Asp Val Gly 20
25 30His Asp Phe Pro Asp Val Gln Phe Asn Trp
Tyr Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Gln Glu Gln Asn Asn Pro Thr 50
55 60Tyr Arg Val Val Ser Ile Leu Ala Ile
Gln His Lys Asp Trp Leu Ser65 70 75
80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn Gln Ala Leu Pro
Ala Pro 85 90 95Val Gln
Lys Thr Ile Ser Lys Pro Thr Gly Gln Pro Arg Glu Pro Gln 100
105 110Val Tyr Val Leu Ala Pro His Pro Asp
Glu Leu Ser Lys Asn Lys Val 115 120
125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro Asp Ile Asp Ile
130 135 140Glu Trp Lys Ser Asn Gly Gln
Pro Glu Pro Glu Thr Lys Tyr Ser Thr145 150
155 160Thr Pro Ala Gln Leu Asp Gly Asp Gly Ser Tyr Phe
Leu Tyr Ser Lys 165 170
175Leu Thr Val Glu Thr Asn Arg Trp Gln Gln Gly Thr Thr Phe Thr Cys
180 185 190Ala Val Met His Glu Ala
Leu His Asn His Tyr Thr Glu Lys Ser Val 195 200
205Ser Lys Ser Pro Gly Lys 210199214PRTArtificial
SequenceAglycosyl variant equine IgG7 Fc, T(64)XMISC_FEATURE(64)..(64)X =
any amino acid except T or S 199Val Gly Pro Ser Val Phe Ile Phe Pro Pro
Lys Pro Lys Asp Val Leu1 5 10
15Met Ile Ser Arg Thr Pro Thr Val Thr Cys Val Val Val Asp Val Gly
20 25 30His Asp Phe Pro Asp Val
Gln Phe Asn Trp Tyr Val Asp Gly Val Glu 35 40
45Thr His Thr Ala Thr Thr Glu Pro Lys Gln Glu Gln Asn Asn
Ser Xaa 50 55 60Tyr Arg Val Val Ser
Ile Leu Ala Ile Gln His Lys Asp Trp Leu Ser65 70
75 80Gly Lys Glu Phe Lys Cys Lys Val Asn Asn
Gln Ala Leu Pro Ala Pro 85 90
95Val Gln Lys Thr Ile Ser Lys Pro Thr Gly Gln Pro Arg Glu Pro Gln
100 105 110Val Tyr Val Leu Ala
Pro His Pro Asp Glu Leu Ser Lys Asn Lys Val 115
120 125Ser Val Thr Cys Leu Val Lys Asp Phe Tyr Pro Pro
Asp Ile Asp Ile 130 135 140Glu Trp Lys
Ser Asn Gly Gln Pro Glu Pro Glu Thr Lys Tyr Ser Thr145
150 155 160Thr Pro Ala Gln Leu Asp Gly
Asp Gly Ser Tyr Phe Leu Tyr Ser Lys 165
170 175Leu Thr Val Glu Thr Asn Arg Trp Gln Gln Gly Thr
Thr Phe Thr Cys 180 185 190Ala
Val Met His Glu Ala Leu His Asn His Tyr Thr Glu Lys Ser Val 195
200 205Ser Lys Ser Pro Gly Lys 210
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